SAFE TRAVEL TIPS Personal safety demands that every attendee take the following precautions: 9 NEVER WALK

ALONE ON CITY STREETS, especially after dark. 9 NEVER WALK OR WAIT ALONE in deserted areas such as public or Conference facility parking lots, hallways, and stairwells. 9 ALWAYS REMOVE your badge when leaving the Conference facility. 9 NEVER accept a ride in an unmarked taxi or shuttle van. 9 NEVER give your hotel room number to strangers. 9 ALWAYS lock up money and valuables. Do not carry them with you to Conference or public facilities. 9 Take your electronic hotel key card home with you, cut it up and discard it. It contains personal information much like a credit card and should be handled accordingly.

47th Annual Drosophila Research Conference PROGRAM ADDENDUM

Friday, March 31
Platform Presenter Changes Cytoskeleton and Cellular Biology Lanier Grand Ballroom A-B 26 - 8:45 am Fragile X mental retardation protein 1 (FMR1) is required for Drosophila cellularization First author O. Papoulas replaced by J. C. Sisson. Gametogenesis and Sex Determination Lanier Grand Ballroom A-B 90 6:00 pm A nuclear hormone receptor that affects spermathecae number and fertility. First author name changed to Anna K. Allen. Organogenesis Ballroom of the Americas D-F 83 - 6:15 pm Rac function in epithelial tube morphogenesis First author Carolyn Pirraglia replaced by Monn Monn Myat. Schedule Change/Addition 2:30 pm FlyMine Demonstration has been changed to 2:00 pm (Level Three, Room 336) 3:304:00 pm Additional FlyMine Demonstration (Level Three, Room 336)

Saturday, April 1
Platform Presentation Cancellation Pattern Formation I Ballroom of the Americas D-F 106 - 10:00 am Regulation of S-phase entry in the developing eye by the bHLH transcription factor, net. Ryan Herbert. Platform Presentation Cancellation and Replacement Technique and Genomics Ballroom of the Americas A-C 99 - 10:15 am Cancelled: BDGP Gene Disruption Project update. Robert W. Levis. (Now poster presentation/board 781C) 99 - 10:15 am Replaced by: The Carnegie protein trap collection: a versatile research resource. Michael Buszczak1, Shelley Paterno1, Julia Bachman1, Dan Lighthouse1, 2, Ben Ohlstein1, Anna K. Allen*1, 2, Todd Nystul1, Tina Tootle1, Erika Matunis3, Terence Murphy1, Stephanie Owen1, Nathathai Srivali1, Megan Kutzer1, Eva Decotto1, James Wilhelm1, Allan Spradling1. (*name changed from Anna Krueger) (Formerly poster presentation/board 781C)

Schedule Change/Addition 2:30 pm FlyMine Demonstration has been changed to 2:00 pm (Level Three, Room 336) 3:304:00 pm Additional FlyMine Demonstration (Level Three, Room 336)

 Poster Presenter/Co-Author Changes Poster # 278A 499C 780B Co-author names, Peter Gallant and Walter Schaffner, removed from abstract Co-author name corrected to Zhigang Jin Jian Jin, substitute presenter for Mark Biggin

Poster Presentations Cancelled Poster # 157C 175C 184C 222B 250C 285B 376C 486B 579B 580C 655C 715C 756B 828B 1st Author Elizabeth Caygill Louise OKeefe Julie Brill Corinna Lehmann Tuba H. Sural Pado Struffi Yu Chen Jochen Trauner Irene Miguel-Aliaga Irene Miguel-Aliaga Claire Hinkley Bruce Bryan Anne D. Plessis Paymaan Jafar-Nejad

Additional Exhibitors VIEWPOINT LIFE SCIENCES, INC. 2550 Bates Street, Ste 404 Montreal, QC, H3S 1A7 Canada tel: (514) 343-5003 fax: (514) 343-5023 e-mail: contact@VPLSI.com URL: http://www.VPLSI.com Booth 204

ViewPoint LS design systems for automation of behavior analysis. We can provide you with complete solutions to fit your needs from hardware to software. VideoTrack <http://62.193.209.148/en/content.php?content.8> system measures animal locomotion and behavior. Numeriscope <http://62.193.209.148/en/content.php?content.13> is a smart digital video recorder for life sciences. LabWatcher <http://62.193.209.148/en/content.php?content.16> is an event recorder.

 Additional Exhibitors, continued CLEVER SYS, INC. 11425 Issac Newton Square Suite 202 Reston, VA 20190 tel: (703) 787-6946 fax: (703) 757-7467 e-mail: nzhang@cleversysinc.com URL: www.cleversysinc.com Booth 303

Headquartered in metropolitan DC area, Clever Sys., Inc., (CSI) develops and sells products and services for lab animal behavior analysis, including rodents, Drosophila, zebra fish, primates, etc. CSIs products are built with technologies of next generation, utilizing information of animal full body and body parts, providing measurements of novel behavioral paradigms and new parameters that have never been available before.

______________________________

y Late Poster Abstracts (see complete text of abstracts beginning on page 12): First Author/Presenter Poster # Abstract Title and Co-Authors

CELL DIVISION AND GROWTH CONTROL Tom Hartl 880C The condensin subunit CapH2 ensures faithful separation of male meiotic chromosomes. Tom Hartl, Sarah Sweeney, Paula Campbell, Giovanni Bosco. Dept Molec & Cellular Biol, Univ Arizona, Tucson. A functional genomics screen identifies Mat89Bb as a novel cell cycle regulator. Michael Anderson, Laura Lee. Dept Cell & Developmental Biol, Vanderbilt Univ, Nashville, TN. A role of histone chaperon ASF1 and Tousled-like kinase in cell cycle progression. Maxim Pilyugin1, Yuri Moshkin2, Franois Karch1. 1) University of Geneva, Switzerland; 2) Erasmus University Medical Centre, Netherlands. Misato is required for centrosome separation in both mitosis and male meiosis. Violaine Mottier, Fiammetta Verni, Giovanni Cenci, Maurizio Gatti, Silvia Bonaccorsi. Genetica e Biologia Moleculare, Universita di Roma La sapienza, Roma, Italy. Characterization of ZW10 in Cytokinesis. Alan Wainman, Maria Grazia Giansanti, Michael Goldberg, Maurizio Gatti. Genetica e Biologia moleculare, Universita di Roma La Sapienza, Roma, Italy. Mechanisms and effects of cell death in Minute-induced cell competition. Wei Li, Nicholas E. Baker. Dept. of Molecular Genetics, Albert Einstein College of Medicine, Bronx, NY 10461. The role of the gene white in increase of mosaic clone frequency in D. melanogaster and D. simulans. Roman Sidorov1, Elena Ugnivenko2, Kirill Kirsanov2, Elizabeth M. Khovanova2. 1) Inst Carcinogenesis, Cancer Research Center RAMS, Moscow, Moscow Region, Russian Federation; 2) Koltzov Institute of Developmental Biology RAS, Moscow, Moscow Region, Russian Federation. A Protein Interactin Map Guided Screen for Novel Regulators of the Cell Cycle. Stephen T. Guest1, Maria A. Cypher1, Russell L. Finley Jr.1, 2. 1) Center for Molecular Medicine and Genetics; 2) Department of Biochemistry and Molecular Biology, Wayne State University School of Medicine, 540 East Canfield Ave, Detroit Michigan 48201. The Regulation of the Tumor Suppressor Activity, PTEN by Thioredoxin Controls Cell Number and Cell Size. Zuohe Song1,2, Negin Saghafi1, Marc Brabant1,3, Emmanuelle Meuillet1,2,3. 1) Arizona Cancer Ctr, Univ Arizona, Tucson, AZ; 2) Nutritional Sciences Department, University of Arizona, 1177 E. 4th Street, Tucson, AZ. 85721; 3) Molecular and Cellular Biology Department, University of Arizona, Tucson, AZ. 85724.

Michael Anderson

881A

Maxim Pilyugin

882B

Violaine Mottier

883C

Alan Wainman

884A

Wei Li

885B

Roman Sidorov

886C

Stephen T. Guest

887A

Zuohe Song

888B

CYTOSKELETON AND CELLULAR BIOLOGY Regulation of autophagosome formation by paxillin during nutrient stress and development. Guang-Chao Chen1,3, Sheila Thomas2, Jeffrey Settleman1. 1) MGH Cancer Center, Charlestown, MA; 2) Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA; 3) Inst of Biological Chemistry, Academia Sinica, Taipei, Taiwan. Regulation of Cell Shape During Eye Organogenesis. Douglas Corrigall, Franck Pichaud. MRC Laboratory for Molecular Cell Biology, UCL, London, United Kingdom.

Guang-Chao Chen

889C

Douglas Corrigall

890A

Masyasu Hirano Manish Jaiswal

891B

892C

Ella Palmer 893A Krishanu Ray Florian Ulrich Slobodan Beronja Giovanna Mottola

894B 895C 896A

897B

Neal T. Sweeney

898C

Sang-Chul 899A Nam Julie Tan 900B

The role of Planar Cell Polarity Effector genes in the embryonic epidermis. Masyasu Hirano1, Janie Coe1, Obianuju Dike1, Nan Ren2, Paul Adler2, Simon Collier1. 1) Dept Biological Sci, Marshall Univ, Huntington, WV; 2) Dept. Biology, University of Virginia. Regulation of epithelial cell-cell adhesion during wing development in Drosophila. Manish Jaiswal, Pradip Sinha. Bio Sci & Bioengineering, Indian Institute Technology, Kanpur, India. A role for the Tuberous Sclerosis Complex during photoreceptor apical membrane differentiation. Ella Palmer, Franck Pichaud. Anatomy and Developmental Biology, UCL. MRC LMCB & CBU, Gower Street, London. WC1E 6BT, United Kingdom. Motors and Morphogenesis: Role of Dynein and Dynactin in testicular fusome assembly in Drosophila. Krishanu Ray, Anindya Ghosh-Roy, Rishita Chengede, Madhura Kulkarni. Dept. of Biological Sciences, Tata Inst. Fund. Res. (TIFR), Mumbai, Maharashtra, India. In vivo analysis of Drosophila ventral furrow formation. Florian Ulrich, Eric Wieschaus. Dept Molecular Biol, Princeton Univ, Princeton, NJ. Cadherin-catenin complex regulates plasma membrane localization of the exocyst complex component Sec6. Slobodan Beronja, Ulrich Tepass. Department of Zoology, University of Toronto, ON, Canada. Endocytic control of wing development in Drosophila m. Giovanna Mottola1,2, Eric Marois1, Suzanne Eaton1, Marcos Gonzlez-Gaitn1, Marino Zerial1. 1) Max Planck Institute for Cell Biology and Genetics, Dresden, Germany; 2) Dipartimento di Biochimica e Biotecnologie Mediche, Universita' di Napoli Federico II, Napoli, Italy. Control of Neuronal Morphogenesis and the Delta/Notch Signaling Pathway by Shrub, a Novel Coiled-Coil Protein in Drosophila. Neal T. Sweeney1, 2, Jay E. Brenman3, Yuh Nung Jan2, 4, Fen-Biao Gao1, 2, 5. 1) Gladstone Institute of Neurological Disease at UCSF, San Francisco, CA; 2) Neuroscience Program, UCSF, San Francisco, CA; 3) Department of Cell and Developmental Biology and Neuroscience Center University of North Carolina, Chapel Hill, NC; 4) Department of Physiology and Howard Hughes Medical Institute, UCSF, San Francisco, CA; 5) Department of Neurology, UCSF, San Francisco, CA. Domain-specific early and late function of Dpatj in Drosophila photoreceptor cells. SangChul Nam, Kwang-Wook Choi. Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX 77030. The role of phosphatidylinositol 4-kinase III in development. Julie Tan1,2, Julie Brill1,2. 1) Developmental Biology, Hospital for Sick Children, Toronto, ON, Canada; 2) Molecular and Medical Genetics, University of Toronto, ON, Canada.

GENOME AND CHROMOSOME STRUCTURE Jennifer A. 901C Armstrong Meridith Toth Craig M. Hart Edward Ramos Karen Weiler 902A CHD1: a chromodomain-containing chromatin remodeling factor. Jennifer A. Armstrong, Matthew S. Berger, Jennifer M. Lee, Ivy E. McDaniel, Nick L. Reeves. Joint Science Dept, Claremont Colleges, Claremont, CA. Deciphering chromatin based cellular processes in development using a dominant negative histone acetyltransferase. Meridith Toth, Xianmin Zhu, Neetu Singh, Felice Elefant. Dept Bioscience/Biotechnology, Drexel Univ, Philadelphia, PA. A dominant negative form of the BEAF insulator proteins disrupts chromatin structure. Craig M. Hart, Matthew Gilbert, Swarnava Roy, Yian Yee Tan. Dept Biological Sciences, Louisiana State Univ, Baton Rouge, LA. Identification and Characterization of Gypsy-like Endogenous Insulators in D. melanogaster. Edward Ramos, Kelly Baxter, Victor Corces. Dept of Biology, Johns Hopkins University, Baltimore, MD. Mapping and mutagenesis of the E(var)3-9 locus. Karen Weiler. Dept of Biology, West Virginia University, Morgantown, WV.

903B

904C 905A

Simon Titen Xiao-Feng Zheng

906B 907C

Xiaolin Bi

908A

Transmission of Fragment Chromosomes Lacking An Endogenous Telomere. Simon Titen, Kent Golic. Dept Biol, Univ Utah, Salt Lake City, UT. Characterizing Drosophila telomeres that lack natural transposons. Xiao-Feng Zheng, Yikang Rong. Lab of Molecular Cell Biology, National Cancer Institute, Bethesda, MD. Partially redundant roles of Drosophila ATM and ATR checkpoint kinase for telomere maintenance. Xiaolin Bi1, Deepa Srikanta1, Laura Fanti2, Sergio Pimpinelli2, Yikang Rong1. 1) NCI/NIH, Bethesda, MD; 2) Dipartimento di Genetica e Biologia Molecolare,Universita di Roma"La Sapienza," Rome, Italy.

REGULATION OF GENE EXPRESSION Vincent C. Henrich 909B A mutational analysis of ecdysone receptor and Ultraspiracle interaction in a heterologous cell culture system. Vincent C. Henrich, Josh Beatty, Jenna Callender. Inst. Health, Science, and Soc, University of North Carolina-Greensboro. Disruption of gene function by targeted insertion and inducible RNAi demonstrate nautilus is essential for embryonic myogenesis and viability and impacts founder cell patterning. Qin Wei1, Yikang Rong2, Bruce Patterson1. 1) Laboratory of Biochemistry, NCI/NIH, Bethesda, MD; 2) Laboratory of Cellular and Molecular Biology, NCI/NIH, Bethesda, MD. Eif1A mutations dominantly suppress the eggshell patterning defect caused by activation of the meiotic recombination checkpoint. Martha Klovstad1, Gail Barcelo1, Uri Abdu2, Trudi Schpbach1. 1) HHMI, Molecular Biology Dept., Princeton University, Princeton, NJ; 2) Dept. of Life Sciences, Ben-Gurion University. Regulation of Histone Acetyltransferase during Development. Vijayalakshmi Ramakrishnan. Drexel University, Philadelphia, PA. Functional characterization of the histone acetyltransferase Elp3 during development. Neetu Singh, Felice Elefant. Dept Bioscience/Biotechnology, Drexel University, Philadelphia, PA. Functional analysis of the chromatin regulator protein: TIP60. Xianmin Zhu, Felice Elefant. Dept Bioscience/Biotechnology, Drexel Univ, Philadelphia, PA. Functional analysis of promoter-enhancer interactions at the Drosophila bithorax complex. Omar S. Akbari, Esther Bae, Robert Drewell. Biology, University of Nevada, Reno.

Qin Wei

910C

Martha Klovstad 911A Vijayalakshmi Ramakrishnan Neetu Singh Xianmin Zhu

912B 913C 914A

Omar S. Akbari 915B SIGNAL TRANSDUCTION

Baoxue Ge

916C

Baoxue Ge

917A

Ning Wang

918B

Dong Yan

919C

Drosophila TAB2 is required for the immune activation of JNK and NF-kappaB. Baoxue Ge1, Zi-Heng Zhuang1, Lei Sun1, Ling Kong1, Jun-Hao Hu1, Ming-Can Yu1, Peter Reinach2, Jin-Wu Zang1. 1) Institute of Health Sciences, Shanghai, China; 2) Department of Biological Sciences, SUNY College of Optometry, New York, NY 10036. Regulation of Drosophila p38 activation by specific MAP2 kinase and MAP3 kinase in response to different stimuli. Baoxue Ge1, Zi-Heng Zhuang1, Yuan Zhou1, MingCai Yu1, Neal Silverman2. 1) Institute of Health Sciences, Shanghai, China; 2) Division of Infectious Disease, Department of Medicine, University of Massachusetts Medical School, 364 Plantation St, Worcester MA 01605. Role of PP2A in visual signaling. Ning Wang, Bih-Hwa Shieh. Dept of Pharmacology, Vanderbilt University, Nashville, TN. Drosophila glypican Dally-like is essential for Breathless mediated tracheal development in embryos and wing discs. Dong Yan1,2, Xinhua Lin1,2. 1) Division of Developmental Biology, Cincinnati Children's Hospital, Cincinnati, OH; 2) Molecular and Developmental Biology Graduate Program, University of Cincinnati, OH.

Li Xiao

920A

Kiranmai Kocherlakota

921B

Xiaoyan Zheng

922C

Sekyung Oh

923A

Bernhard Hovemann

924B

The Rac1 and Rac2 C-terminal Regions Interact Differently with dPosh and Ter94. Li Xiao, Douglas Ruden. Environmental Health Sciences, University of Alabama at Birmingham. An investigation of the oligomerization properties of SNS during myoblast fusion. Kiranmai Kocherlakota1,2, Erika Geisbrecht1, Susan Abmayr1. 1) Stowers Institute for Medical Research, 1000 E 50th Street, Kansas City, MO 64110; 2) Cell and Developmental Biology Department, Huck Institute of Life Sciences, Pennsylvania State University, PA 16802. The Ihog cell surface proteins bind and mediate response to the Hedgehog signal. Xiaoyan Zheng2, Shenqin Yao2, Lawrence Lum2, 4, Philip Beachy1, 2, 3. 1) Howard Hughes Medical Institute; 2) Molecular Biology and Genetics, The Johns Hopkins University School of Medicine, Baltimore, MD; 3) Department of Oncology, The Johns Hopkins University School of Medicine, Baltimore, MD; 4) Current address: Department of Cell Biology, Univeristy of Texas Southwestern Medical Center, Dallas. Reconstitution of Hedgehog signaling from exogenously introduced pathway components. Sekyung Oh, Philip A. Beachy. Molecular Biology & Genetics, Howard Hughes Medical Institute, The Johns Hopkins University School of Medicine, Baltimore, MD. Drosophila cysteine peptidase Tan: expression in photoreceptor cells R1-R8 lends support for a putative histamine neurotransmitter / carcinine cycle between photoreceptor terminals and epithelial glia. Bernhard Hovemann1, Stefanie Wagner1, Christiane Heseding1, John True2. 1) Dept Chemistry, Ruhr Univ, Bochum, Germany; 2) Department of Ecology and Evolution, State University of New York, Stony Brook, NY 11794-5245.

PATTERN FORMATION Hyunsuk Suh 925C Identification of a novel gene that is regulated by engrailed and functions in Drosophila embryo segmentation. Hyunsuk Suh, Jeongbin Yim. Biological Sciences, Seoul National University, Seoul, Korea. Kinesin and Dynein are required for anterior-specific bicoid mRNA transport in the Drosophila oocyte. Byeong J. Cha1, William E. Theurkauf2. 1) Dept Cell & Developmental Biol, Vanderbilt Univ Medical Ctr, Nashville, TN; 2) Program in Molecular Medicine, University of Massachusetts Medical School, Worcester. Differentiation and identity of the Drosophila leg muscles: roles of the FGF pathway and homeodomain transcription factor Ladybird. Tariq Maqbool, Cedric Soler, Teresa Jagla, Jean Philippe Da Ponte, Krzysztof Jagla. INSERM.384, Faculty of Medicine, 28Place Henri Dunant, 63000-Clermont-Fd, France. Characterizing the Role of Drosophila LanA in Ommatidial Development. Yu-Chin Lin1,2, Pei-I Tsai1, Cheng-Ting Chien1. 1) Academia Sinica, Institute of Molecular Biology, Taipei, Taiwan; 2) Institute of Neuroscience, National Yang-Ming University, Taipei, Taiwan. The Control of Ommatidial Death in the Peripehry of the Fly Eye. Andrew Tomlinson, Hui-Ying Lim. Dept Genetics & Development, Columbia Univ, New York, NY. Functional characterization of CG32062, the Drosophila homologue of human Ataxin-2 Binding Protein. Nagarajan Usha, LS Shashidhara. Centre for Cellular and Molecular Biology, Hyderabad, AP, India.

Byeong J. Cha 926A

Tariq Maqbool 927B

Yu-Chin Lin Andrew Tomlinson Nagarajan Usha

928C

929A 930B

GAMETOGENESIS AND SEX DETERMINATION Yu Chen 931C oogenesis. Yu Chen1, 2, Trudi Schpbach1, 2. 1) Howard Houghes Medical Institute; 2) Department of Molecular Biology, Princeton University.

Cutoff, a potential exoribonuclease associated protein, is required for Drosophila

Elizabeth Eldon Horacio M. Frydman

932A

933B

Jason Burgess 934C

Thomas Fellner

935A

A role for the Toll-like receptor, 18-Wheeler, in ovarian follicle cell migration. Elizabeth Eldon, Dominic Siler, Cassandra Kleve, Marie Alpuerto. Dept Biological Sci, California State Univ, Long Beach. Wolbachia preferentially populate the somatic stem cell niche. Horacio M. Frydman1,2, Jennifer Li1,2, Drew Robson1,2, Eric Wieschaus1,2. 1) Howard Hughes Medical Institute; 2) Dept Molecular Biol, HHMI, Princeton Univ, Princeton, NJ. Phosphatidylinositol 4-kinase type II is required for spermatogenesis in Drosophila. Jason Burgess, Ho-Chun Wei, Gordon Polevoy, Julie A. Brill. Dept Developmental Biol, Rm 13-401-M, Hospital for Sick Children, Toronto, ON, Canada, M5G 1L7. lucky luke (luke), a novel gene with a potential function in asymmetric stem cell division. Thomas Fellner1, Allyson Spence1, Cordula Schulz2. 1) Department of Developmental Biology, Stanford University School of Medicine, Stanford, CA; 2) Cold Spring Harbor Laboratories, Cold Spring Harbor, NY.

ORGANOGENESIS Regulation of Drosophila Friend of GATA gene, u-shaped, during hematopoiesis: A direct role for Serpent and Lozenge. Selen Muratoglu1, Betsy Garratt1, Kristy Hyman2, Kathleen Gajewski2, Robert A. Schulz2, Nancy Fossett1. 1) Center for Vascular and Inflammatory Diseases and the Department of Pathology, University of Maryland School of Medicine, Baltimore, MD 21201; 2) Department of Biochemistry and Molecular Biology, Graduate Program in Genes & Development, The University of Texas M. D. Anderson Cancer Center, Houston, TX 77030. Expression and mutual repression of tinman and the Dorsocross genes establishes diversified cardiac cell identities in the dorsal vessel. Ingolf Reim1, Stphane Zaffran2, Manfred Frasch1. 1) Mol., Cell & Dev. Biology, Mount Sinai School of Medicine, New York, NY; 2) Dev. Biology Inst. of Marseille-Luminy, Marseille, France. Positive and Negative Control of Muscle Differentiation. Michael Taylor, David Liotta. Sch Biosciences, Cardiff Univ, Cardiff, United Kingdom.

Selen Muratoglu

936B

Ingolf Reim

937C

Michael Taylor 938A

NEUROGENETICS AND NEURAL DEVELOPMENT The F-actin/Microtubule crosslinker Shot is a platform for Krasavietz-mediated translational regulation of midline axon repulsion. Seungbok Lee1, Seongsoo Lee1, Peter Kolodziej2. 1) School of Dentistry, Seoul National Univ, Seoul, Korea; 2) Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN. Highly ordered assembly of retinal axons and their synaptic partners is regulated by Hedgehog/Single-minded in Drosophila visual system. Daiki Umetsu, Satoshi Murakami, Makoto Sato, Tetsuya Tabata. Dept Cell Biol, IMCB Univ of Tokyo, Japan. Screening of novel genes related to neuronal apoptosis mediated by dAPP-BP1 pathway in D. melanogaster. Songhee Kim, Kiyoung Kim, Joohee Lee, Jongwoo Ryu, Jeongbin Yim. School of Biological Sciences, Seoul National University, Seoul, Korea. Confocal analysis of the Drosophila mutant small mushroom bodies (smu) reveals an abnormal cell number and axonal pattern. Brian Dunkelberger, Christine Serway, Steven de Belle. University of Nevada, Las Vegas, 89154-4004.

Seungbok Lee 939B

Daiki Umetsu 940C

Songhee Kim 941A Brian 942B Dunkelberger

Michelle 943C Brooks-Pigott

Analysis of the role of Drosophila Phosphotyrosyl phosphatase activator (D-Ptpa) in peripheral nervous system development. Michelle Brooks-Pigott1,2,3, Eddy Van Daele4, Veerle Janssens5, Christine Van Hoof5, Jozef Goris5, Koen Norga1,2,6, Patrick Callaerts1,2,3. 1) Laboratory of Developmental Genetics, Flanders Institute of Biotechnology (VIB); 2) Center for Human Genetics, University of Leuven, Leuven, BE; 3) University of Houston, Department of Biology and Biochemistry, Houston, Texas (former affiliation); 4) University of Sussex, School of Biological Sciences, Brighton, UK; 5) University of Leuven, Afdeling Biochemie, Faculteit Geneeskunde, Departement Moleculaire Celbiologie, Leuven, BE; 6) Childrens Hospital, University of Leuven, Leuven, BE.

NEURAL PHYSIOLOGY AND BEHAVIOR Lixian Zhong 944A The Drosophila Mutant oblivious Is Defective In Nociception. Lixian Zhong4, W. Daniel Tracey1,2,3. 1) Dept. of Anesthesiology; 2) Dept. of Cell Biology; 3) Dept. of Neurobiology; 4) Dept. of Pharmacology, Duke University, Durham, NC. A Sleep-promoting Role for the Drosophila Serotonin Receptor 1A. Quan Yuan1, William Joiner2, Amita Sehgal1, 2. 1) Howard Hughes Medical Institute; 2) Center for Sleep and Respiratory Neurobiology, University of Pennsylvania, School of Medicine, Philadelphia, PA. Classical reward conditioning in D. melanogaster. Kyung-An Han, Young-Cho Kim, HyunGwan Lee. Department of Biology, Pennsylvania State Univ, University Park. Drosophila Dorsal Paired Medial neurons provide a general mechanism for prolonging memory. Michael J. Krashes, Alex C. Keene, Jessica A. Bernard, Scott Waddell. Neurobiology, Univ of Massachusetts Medical School, Worcester. Multiple memories of Drosophila sugar reward learning. Andreas S. Thum, Martin Heisenberg, Hiromu Tanimoto. Department of Genetics and Neurobiology, University of Wrzburg, Biozentrum am Hubland, Germany. Protein Phosphatase 1 Regulates the Stability of Drosophila Circadian Clock Proteins. Yanshan Fang, Sriram Sathyanarayanan, Amita Sehgal. Howard Hughes Medical Institute, Department of Neuroscience, University of Pennsylvania School of Medicine, Philadelphia, PA 19104. FOXO and insulin signaling regulate the sensitivity of circadian clock to oxidative stress. Xiangzhong Zheng, Amita Sehgal. Dept. Neurosci, 232 Stemmler Hall, Univ. Pennsylvania Med. Sch./HHMI, Philadelphia, PA 19104.

Quan Yuan Kyung-An Han Michael J. Krashes Andreas S. Thum Yanshan Fang Xiangzhong Zheng

945B

946C 947A

948B

949C

950A

EVOLUTION AND QUANTITATIVE GENETICS Margarita Ramos 951B Evolution Through the Eyes of a Fly. Margarita Ramos, Alistair McGregor, David Stern, Peter Grant. Ecology and Evolutionary Biology, Princeton University, Princeton, NJ.

IMMUNE SYSTEM AND CELL DEATH Collier/Knot is required for PSC formation and control of blood cell homeostasis. Alain Vincent1, Joanna Krezmien1, Rami Makki1, Laurence Dubois1, Jean Michel Ubeda2, Marie Meister2, Michle Crozatier1. 1) Centre de Biologie du Dveloppement, CNRS/Universit, Toulouse, France; 2) Institut de Biologie Molculaire et Cellulaire, CNRS, Strasbourg, France.

Alain Vincent 952C

Lihui Wang

953A

Julien Colombani

954B

Katherine A. 955C Taylor

Establishing the molecular events during pattern recognition of Gram-positive ain Drosophila innate immunity. Lihui Wang1, Alexander Weber2, Sergio Filipe3, s Gay2, Petros Ligoxygakis1. 1) Genetics Unit, Department of Biochemistry, ity of Oxford, South Parks Rd OX1 3QU, Oxford UK; 2) Department of mistry University of Cambridge, 80 Tennis Court Road, CB2 1GA Cambridge UK; uto de Tecnologia Qumica e Biolgica / Universidade Nova de Lisboa, Av. da ca (EAN), Apartado 127, 2781-901 Oeiras, Portugal. Dmp53 activates the Hippo pathway to promote cell death in response to DNA damage. Julien Colombani, Cedric Polesello, Filipe Josue, Nicolas Tapon. Apoptosis and Proliferation Control Lab, Cancer Research UK, London Institute, United Kingdom. Alteration of gravitational force affects the immune response. Katherine A. Taylor1, Alexander T. Kloehn1, Patrick M. Fuller2, Kathleen M. Beckingham3, Charles A. Fuller2, Deborah A. Kimbrell1. 1) Molecular & Cellular Biology, University of California, Davis, CA; 2) Neurobiology, Physiology & Behavior, University of California, Davis; 3) Dept. Biochemistry and Cell Biology, Rice University, Houston, TX.

TECHNIQUES AND GENOMICS An efficient genetic screen in Drosophila to identify nuclear-encoded genes with mitochondrial function. Albert Cespedes1,2, T. S. Vivian Liao1,2, Gerald B. Call1,2, Preeta Guptan1, Jamie Marshall1, Edward Owusu-Ansah1, Sudip Mandal1, Q. Angela Fang1, Kevin Yackle1, Gelsey L. Goodstein1, William Kim1, Utpal Banerjee1. 1) Mol., Cell and Dev. Biology, UCLA, Los Angeles, CA; 2) These authors contributed equally to this work. Insect Sequencing at the Baylor College of Medicine Human Genome Sequencing Center. Stephen Richards1, Yue Liu1, Kim C. Worley1, Erica Sodergren1, Steven E. Scherer1, Catherine M. Rives1, Susan Brown2, Richard Beeman3, John H. Warren4, Donna M. Muzny1, George Weinstock1, Richard A. Gibbs1. 1) Human Genome Sequencing Center, Baylor College of Medicine, Houston TX; 2) Division of Biology, Kansas State University, Manhattan, KA; 3) USDA, Biological Research Unit, 1515 College Ave, Manhattan, KA; 4) Department of Biology, University of Rochester, NY. Use Drosophila as A Model System to Identify the Function of Human Genes. Yung-Heng Chang1,2, Y. Henry Sun2. 1) Graduate Institute of Life Sciences, National Defense Medical Center, Taipei 114, Taiwan, Republic of China; 2) Institute of Molecular Biology, Academia Sinica, Taipei 115, Taiwan, Republic of China.

Albert Cespedes

956A

Stephen Richards

957B

Yung-Heng Chang

958C

DROSOPHILA MODELS OF HUMAN DISEASES An inducible Drosophila cell model to study polyglutamine disease. Wing Man Chan1,2, Pang Chui Shaw2, Edwin HY Chan1,2. 1) Laboratory of Drosophila Research, The Chinese University of Hong Kong, HKSAR, China; 2) Molecular Biotechnology Program, Department of Biochemistry, The Chinese University of Hong Kong, HKSAR, China.

Wing Man Chan

959A

10

Lasbleiz Christelle Allyson Spence

960B

961C

Kyu-Sun Lee 962A

A New Drosophila model for SCA7 polyglutamine disease demonstrates reversible city and allows quantitative screening for phenotypic modifiers. Lasbleiz Christelle1, Vronique1, Latouche Morwena2, Brice Alexis2, Stevanin Govanni2, Tricoire Herv1. 1) cques Monod, Paris, France; 2) Institut des Neurosciences, Paris, France. D. melanogaster as a model of mammalian hematopoiesis: characterization of the spaghetti mutant. Allyson Spence, Thomas Fellner, Margaret Fuller. Department of Developmental Biology, Stanford University School of Medicine, Stanford. Over-expression of JNK signaling rescues electromagnetic stress induced lethality in Drosophila. Kyu-Sun Lee1, Jang-Yul Kwak2, Dong-Seok Lee3, Kweon Yu1. 1) Development & Differentitation, KRIBB, Daejeon, KOREA; 2) Insect Resources Laboratory, KRIBB, Daejeon, KOREA; 3) Laboratory of Human Genomics, KRIBB, Daejeon, KOREA.

PHYSIOLOGY AND AGING William Staatz 963B Regulation of SOD by LAMMER kinases. William Staatz, Sarah Wilkinson, Emmanuelle Meuillet. Arizona Cancer Ctr, Univ Arizona, Tucson.

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47th Annual Drosophila Research Conference Late Poster Abstracts

CELL DIVISION AND GROWTH CONTROL 880C The condensin subunit CapH2 ensures faithful separation of male meiotic chromosomes. Tom Hartl, Sarah Sweeney, Paula Campbell, Giovanni Bosco. Dept Molec & Cellular Biol, Univ Arizona, Tucson, AZ. Condensins I and II are multi-subunit protein complexes that are implicated in chromosome condensation and faithful chromosome separation each cell cycle. Mutations exist in Drosophila condensin I subunits gluon/SMC4, CapG, and barren/CapH that lead to extensive failure in mitotic chromosome separation in embryonic and/or larval brain cell divisions. We have isolated four mutant alleles of the condensin II subunit CapH2, three of which are specifically male sterile and one weaker allele that causes male specific non-disjunction. Cytology of primary spermatocytes from CapH2 mutant males clearly shows a high degree of chromatin disorganization and upon anaphase, chromosomes do not become completely separated while migrating to the poles. Although male meiosis is disrupted in CapH2 mutants, females may have no significant consequence as fertility is only subtly altered and non-disjunction has yet to be observed. Females do have one striking phenotype, the chromosomes in their polyploid nurse cell nuclei become highly polytene throughout the life of the nurse cell. In Xenopus and human tissue culture cells it has been shown that a distinct condensin complex exists that is made up of non-SMC subunits CapD3, CapG2, and CapH2. It is likely that CapH2 and CapD3 also act together in flies as part of a condesin II complex because like CapH2 mutants, flies homozygous for a CapD3 mutant allele are also male sterile and carry polytene chromosomes in their nurse cell nuclei. We propose that CapH2 and CapD3 act as part of a Drosophila condensin II complex that functions in male meiosis to mediate disjoining of chromosomes upon anaphase onset. In addition, this complex is necessary to resolve polytene chromosome attachments and/or prevent their formation during nurse cell development. 881A A functional genomics screen identifies Mat89Bb as a novel cell cycle regulator. Michael Anderson, Laura Lee. Dept Cell & Developmental Biol, Vanderbilt Univ, Nashville, TN. Early embryonic cell cycles in Drosophila consist of rapidly alternating rounds of DNA replication (S phase) and mitosis (M phase) without intervening gaps (G1 and G2). Driven by large maternal pools of RNA and protein, these streamlined cell cycles are regulated by posttranscriptional mechanisms. Three Drosophila genes, pan gu (png), plutonium (plu), and giant nuclei (gnu), play crucial roles in coordinating these rapid S-M cycles. Mutation of any of these three genes disrupts proper coupling between S phase and mitosis, resulting in DNA replication in the absence of mitosis with the accumulation of polyploid nuclei in embryos from homozygous mutant females. This giant nuclei phenotype can be explained, at least in part, by decreased levels of mitotic cyclins in mutant embryos. The png gene encodes a serine/threonine kinase, and plu and gnu encode novel proteins. We have previously shown that proteins encoded by these three genes physically interact to form an active PNG kinase complex. We previously performed a genome-scale biochemical screen for PNG kinase substrates that led to the identification of Mat89Bb, a novel, maternally expressed protein of unknown function. Knockdown of Mat89Bb function in Drosophila embryos by RNAi resulted in a weak giant nuclei phenotype, and knockdown of the Mat89Bb homolog in Xenopus embryos by morpholinos similarly resulted in both cell cycle defects (polyploidy) and developmental arrest. Furthermore, we have also shown that human Mat89Bb is required for somatic cell cycle regulation. These findings have revealed that Mat89Bb plays a critical, conserved role in regulating both the cell cycle and development. Current efforts are directed towards the identification and characterization of Drosophila Mat89Bb mutant lines. A careful phenotypic analysis of these mutants will likely have relevance for our understanding of Mat89Bbs cell-cycle and developmental functions in Drosophila as well as higher organisms. 882B A role of histone chaperon ASF1 and Tousled-like kinase in cell cycle progression. Maxim Pilyugin1, Yuri Moshkin2, Franois Karch1. 1) University of Geneva, Switzerland; 2) Erasmus University Medical Centre, Netherlands. ASF1 is highly conserved histone chaperon. We have investigated the biological function of ASF1 in control of chromatin dynamics and cell cycle progression using Drosophila as a model organism. We identified two mutations in asf1 gene that appeared to suppress heterochromatin-mediated silencing, demonstrating a role for ASF1 in the proper assembly of silenced chromatin. We have performed genetic interactions experiments that led us to discover that chromatin assembly factor ASF1 cooperates in vivo with the Brahma ATP-dependent chromatin remodeling machinery. We also performed ASF1 RNAi knockdown study in Drosophila cell culture. The global changes of gene

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47th Annual Drosophila Research Conference Late Poster Abstracts expression in ASF1 depleted cells were analyzed mRNA hybridization on Affymetrix GeneChip array and compared to the changes caused by depletion of BRM. Statistically significant correlation of the gene expression changes has been found in the two sets of data. We showed that ASF1 is a phosphorylation target of cell cycle regulated Tousled-like kinase (TLK). The analysis of DNA content in TLK RNAi treated cells implies that the reduction of TLK activity slowdown the transition from G1 to S phase. Our data suggest as well that TLK activity affects ASF1 protein stability providing a mechanism for ASF1 function regulation. As a histone chaperone, ASF1 may influence a chromatin structure of genes and affect their expression. Our results suggest that high level of ASF1 may exacerbate regulation by E2F in vivo. The comparison of gene expression changes in Drosophila cell depleted of E2F complex components, ASF1, TLK, and BRM implies the role for ASF1 and BRM in transcriptional regulation of the genes controlled by E2F and suggests that ASF1 acts as a co-activator for E2F1 regulated genes with assistance of BRM. Therefore, both in vivo and in vitro experiments suggest that E2F and ASF1 are acting together in the regulation of E2F target genes via the SWI/SNF complex. 883C Misato is required for centrosome separation in both mitosis and male meiosis. Violaine Mottier, Fiammetta Verni, Giovanni Cenci, Maurizio Gatti, Silvia Bonaccorsi. Genetica e Biologia Moleculare, Universita di Roma La sapienza, Roma, ITALY. The Drosophila misato (mst) gene encodes a protein that contains motifs shared among tubulins, as well as a motif related to the myosin heavy chain. Previous studies showed that the mst mutant alleles mstLB20 and mstC27 are late lethal and exhibit frequent polyploid cells in larval brains (Miklos et al., 1997). However, the precise function of the Misato protein remains unknown. Sequencing of the mst gene in the LB20 and C27 mutant alleles shows that both mutations lead to truncated proteins. Examination of mst mutant brains reveals that most mitotic spindles (70%) are monopolar, suggesting a defect in centrosome separation. In addition, these monopolar spindles display shorter and fewer microtubules than wild type spindles. Nevertheless, recruitment of the centrosomal proteins CNN, -tubulin, D-TACC, CP190, CP60, DGRIP91, DPLP is not affected in mst mutants. mst mutants do not show major defects during the first meiotic division but display frequent spindles (67%) with unseparated centrosomes during the second division. These findings can be explained considering that the first meiotic division is considerably longer than the second, giving the centrosomes enough time to separate from each other. Using a monoclonal antibody raised against Misato, we find that this protein localizes at the spermatid basal body, which is a centrosome-derived structure. However, no centrosomal localization is observed in dividing cells. We are generating flies expressing a Misato-GFP fusion protein to determine whether Mst is enriched at mitotic and meiotic centrosomes. 884A Characterisation of ZW10 in Cytokinesis. Alan Wainman, Maria Grazia Giansanti, Michael Goldberg, Maurizio Gatti. Genetica e Biologia moleculare, Universita di Roma La Sapienza, Roma, ITALY. ZW10, along with the other member of the Rod, Zwilch, Zw10 (RZZ) complex, are required for the proper functioning of the mitotic spindle checkpoint. In addition to this well characterized role, the male meiotic divisions of zw10 mutants display defects in cytokinesis, whereas rod and zwilch mutants show no cytokinesis defects. Here we demonstrate that all three RZZ components concentrate within the spindle envelope remnant during late anaphase, and during telophase aggregate to form a band across the mid-zone of the spindle envelope. We also present a phenotypic analysis of zw10 mutants demonstrating that cytokinesis failure is similar to that observed in giotto, four wheel drive and four way stop mutants. These results suggest a role for ZW10 in mediating membrane dynamics during cytokinesis, consistent with recent findings indicating the human homologue of ZW10 is involved in Golgi trafficking. 885B Mechanisms and effects of cell death in Minute-induced cell competition. Wei Li, Nicholas E. Baker. Dept. of Molecular genetics, Albert Einstein College of Medicine, Bronx, NY 10461. Cell competition has been reported to play important roles in growth regulation and organ size control. To investigate the mechanisms and effects of cell death during cell competition, we examined extensive interfaces of competing populations when +/+ cells were generated by mitotic recombination in M/+ backgrounds. We found that cell death in cell competition happens only at specific locations and that such characteristic cell death is required for cell competition. We focused on such site specific cell death to study the roles of cell intermingling and Dpp signaling in competition. Both are affected by the cell death. The enhanced growth that cell competition induces in

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47th Annual Drosophila Research Conference Late Poster Abstracts the +/+ cells was also studied. We found that such enhanced growth depended on death of M/+ cells and differed mechanistically from compensatory cell proliferation that has been reported in other situations. We are currently studying genes that are required for cell competition and the possible role of phagocytosis in the process. 886C The role of the gene white in increase of mosaic clone frequency in D. melanogaster and D. simulans. Roman Sidorov1, Elena Ugnivenko2, Kirill Kirsanov2, Elizabeth M. Khovanova2. 1) Inst Carcinogenesis, Cancer Research Center RAMS, Moscow, Moscow Region, RUSSIAN FEDERATION; 2) Koltzov Institute of Developmental Biology RAS, Moscow, Moscow Region, RUSSIAN FEDERATION. In flies heterozygous for white and other X-linked markers the marker linked to the white mutation in cis-, possesses higher mosaic clone frequency than the marker in trans-. E.g., in y + + / + w sn heterozygotes the singed clone frequency is higher than the frequency of the twin yellow clones. This effect is true in two Drosophila species, D. simulans and D. melanogaster, for both spontaneous and induced by various agents (-irradiation, chemical mutagens, H+-factor of genetic instability) somatic mosaicism. The effect is more obvious for small clones. It depends upon the irradiation dose and developmental stage at which the mutagen was applied. 887A A Protein Interactin Map Guided Screen for Novel Regulators of the Cell Cycle. Stephen T. Guest1, Maria A. Cypher1, Russell L. Finley Jr.1, 2. 1) Center for Molecular Medicine and Genetics; 2) Department of Biochemistry and Molecular Biology, Wayne State University School of Medicine, 540 East Canfield Ave, Detroit Michigan 48201, USA. Large-scale protein interaction maps (PIMs) have now been generated for several model organisms including Drosophila melanogaster.These maps contain many previously identified interactions between members of characterized protein complexes and signaling pathways. The maps also reveal a vast number of novel interactions, including many involving uncharacterized proteins. These interactions illuminate potential novel players in critical cellular processes. We have performed network analysis of the currently available PIM data for Drosophila in search of interactions that identify potential novel regulators of the cell cycle. Our analysis has identified two high confidence subsets of proteins that are involved in novel interactions with known cell cycle regulators. Proteins in the first subset are involved in novel interactions with members of the cyclin dependent kinase family, while proteins in the second subset are involved in novel interactions with members of the SCF ubiquitin ligase complex. To identify a role for these proteins in cell cycle progression, we reduced their expression in cultured Drosophila cells by RNA interference in a 96-well format and looked for cell cycle defects using flow cytometry. As a result of this approach, we have identified several novel regulators of cell cycle progression. Our results validate the use of PIM data in guiding discovery of novel regulators of key cellular processes. *S. Guest and M. Cypher contributed equally to this work. 888B The Regulation of the Tumor Suppressor Activity, PTEN by Thioredoxin Controls Cell Number and Cell Size. Zuohe Song1,2, Negin Saghafi1, Marc Brabant1,3, Emmanuelle Meuillet1,2,3. 1) Arizona Cancer Ctr, Univ Arizona, Tucson, AZ; 2) Nutritional Sciences Department, University of Arizona, 1177 E. 4th Street, Tucson, AZ. 85721, USA; 3) Molecular and Cellular Biology Department, University of Arizona, Tucson, AZ. 85724, USA. Human Thioredoxin-1 (hTrx-1) is a redox protein with a molecular weight of 12 kDa, which contains the two catalytic site cysteine residues -Trp- Cys32-Gly-Pro-Cys35-Lys found in all thioredoxin proteins. hTrx-1 has been reported to play an important role in cell growth, apoptosis, and cancer patient prognosis. Recently, we have demonstrated that hTrx-1 binds to the C2 domain of hPTEN in a redox dependent manner. This binding leads to the inhibition of PTEN lipid phosphatase activity in mammalian systems (1). In this study, we show that overexpression of hTrx-1 in Drosophila Melanogaster promotes cell proliferation during eye development as measured by eye size and ommatidia size. Furthermore, hTrx-1 can rescue the small eye phenotype induced by the overexpression of the tumor suppressor gene PTEN. We demonstrate that hTrx-1 rescue of the PTEN overexpression eye size phenotype requires cysteine218 in the C2 domain of PTEN. In addition, we show that hTrx-1 expression results in increased Akt phosphorylation supporting our conclusion that he hTrx-1 induced eye size increase results from the inhibition of the PTENs PtdIns-3-phosphatase activity. Our study confirms the redox regulation of PTEN through disulfide bond formation with the hTrx-1 in Drosophila and suggests a conserved mechanisms for Thioredoxins and their interactions with the PtdIns-3-kinase signaling pathway in man and the fruit fly.

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47th Annual Drosophila Research Conference Late Poster Abstracts CYTOSKELETON AND CELLULAR BIOLOGY 889C Regulation of autophagosome formation by paxillin during nutrient stress and development. Guang-Chao Chen1,3, Sheila Thomas2, Jeffrey Settleman1. 1) MGH Cancer Center, Charlestown, MA; 2) Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA; 3) Inst of Biological Chemistry, Academia Sinica, Taipei, TAIWAN. We have previously shown that Drosophila paxillin functions in regulation of Rho GTPases in tissue morphogenesis (Chen et al. 2005, Mol Cell Biol). In order to further elucidate its role in development, we undertook a misexpression screen to isolate new functional partners of paxillin. Wing-specific expression of paxillin induces blistered wings, and this phenotype was used as the basis for a modifier screen of ~1500 EP lines. One of the candidates identified corresponds to the serine/threonine kinase Center divider (Cdi, EP3319), a homolog of Drosophila LIM kinase, which is involved in the regulation of the actin cytoskeleton via cofilin. Furthermore, the insertion EP(3)3348 in the 5UTR of Drosophila Ulk was isolated as a strong suppressor of the paxillin-induced wing blistering phenotype, but has no effect on the Dlimk-induced wing phenotype. The dUlk mutant generated by the P element excision of EP3348 fails to suppress the paxillin-induced wing phenotype, suggesting that dUlk may be a specific interactor with paxillin. Ulk is involved in autophagosome formation and axon outgrowth. In Drosophila, ecdysone receptor signaling activates programmed autophagy. Interestingly, ecdysone receptor mutations suppress paxillin induced wing phenotype. We are currently investigating the role of paxillin in autophagosome formation and neuronal differentiation in development. 890A Regulation of Cell Shape During Eye Organogenesis. Douglas Corrigall, Franck Pichaud. MRC Laboratory for Molecular Cell Biology, UCL, London, UNITED KINGDOM. Control of cell shape has been shown in many diverse metazoan phyla to be crucial to the proper development of tissues and organs. This cell response is often polarized and by definition involves rearrangement of the Factin/microtubule cytoskeleton downstream of patterning cues. To address this problem, we are using the striking apical cell constriction in the fly eye that is associated with the morphogenetic furrow (MF) downstream of the conserved Dpp and Hh pathways. We have initiated a pilot screen using available FRT lines corresponding to a subset of the cytoskeletal-related factors identified as regulators of cell shape in an S2 cell based system (Kiger et al. 2003). This screen demonstrates that i) we can detect robust phenotypes affecting the MF cell response, and ii), only a minority of these factors might be relevant in this process. In particular we will present evidence that loss-offunction clones of the cofilin-phosphatase slingshot (ssh) give a striking phenotype in the MF, in which hyperaccumulation of F-actin is consistent with a failure to inhibit cofilin activity and results in loss of the MF cell response and prevention of proper neuronal differentiation. 891B The role of Planar Cell Polarity Effector genes in the embryonic epidermis. Masyasu Hirano1, Janie Coe1, Obianuju Dike1, Nan Ren2, Paul Adler2, Simon Collier1. 1) Dept Biological Sci, Marshall Univ, Huntington, WV; 2) Dept. Biology, University of Virginia, VA. During late embryogenesis the epidermal epithelium becomes patterned with polarized structures. Rows of denticles appear on the ventral side and the dorsal surface becomes almost entirely covered in cell hairs. The Planar Cell Polarity (PCP) Effector genes, fritz, fuzzy, inturned and multiple wing hairs are required for the correct spacing, orientation and number of these structures. Our analysis of PCP Effector gene mutant phenotypes reveals that these genes control epithelial cell shape, cell polarity and the number of denticle or hairs produce per cell. Changes in F-actin distribution in PCP Effector mutants and the requirement for small GTPase function suggests that the regulation of actin cytoskeleton is critical for embryonic epidermal patterning. Significantly, a Fritz-GFP fusion protein localizes with F-actin throughout the developing epidermal epithelium. We propose a three-step model for epithelial patterning that requires the coordinated control of cell shape, cell polarity and the formation of cell structures. All three steps are mediated by the actin cytoskeleton under the regulation of the PCP Effector genes. 892C Regulation of epithelial cell-cell adhesion during wing development in Drosophila. Manish Jaiswal, Pradip Sinha. Bio Sci & Bioengineering, Indian Institute Technology, Kanpur, INDIA. In a developing organ, intercellular adhesive interactions are involved in sorting and aggregation of cells. Expression and distribution of cell adhesion proteins involved in these interactions are often regulated by instructive cues from the signaling pathways involved in pattern formation and growth. Cell adhesion proteins in turn also coordinate cell-cell communication. A tight coordination between cell-cell adhesion and growth/pattern regulation 15

47th Annual Drosophila Research Conference Late Poster Abstracts thus determines the characteristic size and shape of an organ. In Drosophila, the primordia of the adult appendages i.e., imaginal discs, offer an elegant model to investigate these cellular and developmental mechanisms. Here, we show that the Wg/Wnt and Dpp signaling that regulates growth and patterning of wing imaginal disc also regulates cell-cell adhesion. Both Wg and Dpp signaling provides cues for developmental regulation of DE-Cadherin (DE-Cad) which codes for a homophilic cell adhesion protein. Our results reveal that the Wg signaling mediated regulation of cell adhesion is a function of its transcriptional output rather than a direct interaction of Wg signaling induced elevated Arm/-catenin pool with that of DE-cadherin at the adherence junction. We further show that Fat (Ft), a large cadherin-like protein involved in heterophilic cell-cell adhesion, genetically intersects with the Wg pathway inhibits DE-Cad transcription. The regulation of spatial pattern of cell-cell adhesion and remodel the epithelial cell-cell junctions in the growing wing imaginal disc that determines the final shape of the adult wing. 893A A role for the Tuberous Sclerosis Complex during photoreceptor apical membrane differentiation. Ella Palmer, Franck Pichaud. Anatomy and Developmental Biology, UCL. MRC LMCB & CBU, Gower Street, London. WC1E 6BT. UNITED KINGDOM. The vast majority of cancer in humans originate from epithelia. In order to better understand the nature of the cytoskeletal and polarity instabilities that could lead to tumorogenesis we are using the genetically amenable developing photoreceptor as a model epithelial cell. Completion of the Drosophila genome has led to the identification of approximately 1000 genes involved in cytoskeleton dynamics and these genes are being screened using FRT chromosomes carrying the mutations of interest and in vivo RNAi. We report here the results from a pilot recessive screen that was performed on the right arm of the third chromosome. Among the genes that were examined, we have identified Tuberous Sclerosis Complex 2 (TSC2), an important regulator of cell growth, as a key factor for apical membrane morphogenesis. In absence of this gene function, the morphogenesis of the apical light gathering organelle of the photoreceptor is strongly impaired suggesting that the TSC1 and TSC2 genes play a role in regulating this differentiation process in photoreceptors. 894B Motors and Morphogenesis: Role of Dynein and Dynactin in testicular fusome assembly in Drosophila. Krishanu Ray, Anindya Ghosh-Roy, Rishita Chengede, Madhura Kulkarni. Dept. of Biological Sciences, Tata Inst. Fund. Res. (TIFR), Mumbai, Maharashtra, INDIA. Fusomes are tubulovesicular organelles present in the germ line cysts of holometabolous insects. Studies in Drosophila ovary established that it is rich in spectrin, adducin (Hts) and other spectrin-associated proteins, and it is connected to the endoplasmic reticulum through rapid exchange of membrane. Fusomes contain the microtubuleorganizing center (MTOC) and determine the asymmetric cell divisions as well as the oocyte cell fate in the ovary. In comparison, very little is known about the composition and role of fusome in male germ-line cysts, which differentiates symmetrically to produce sixty-four identical sperm. We find that the Dynein Light Chain 1 (DLC1), along with spectrin and adducin, is enriched in testicular fusomes and loss of the protein in ddlc1 hemizygous testes affect the fusome assembly. In addition, the results of a genetic interaction analyses suggest that both the intermediate chain subunit (IC74, sw1) of the cytoplasmic dynein and the P150dynactin (Glued) are required for the testicular fusome assembly. They are required for DLC1 localization to the organelle. Altogether, these results have provided a new insight into the mechanism of fusome assembly in particular and the endomembrane organization in general in Drosophila. 895C

In vivo analysis of Drosophila ventral furrow formation. Florian Ulrich, Eric Wieschaus. Dept Molecular Biol,
Princeton Univ, Princeton, NJ. During Drosophila ventral furrow (VF) formation, mesodermal precursors separate from the prospective neurectoderm through a sequence of rapid cell shape changes, including apical flattening and constriction, cell elongation and subsequent shortening, leading to the internalization of the cells along the anterior-posterior axis. VF formation is induced by the patterning genes dorsal and twist that regulate the expression of folded gastrulation (fog). The secreted Fog protein localizes myosin II towards the apical side of prospective mesodermal cells via a pathway involving RhoGEF2 and Drosophila Rho-associated kinase (Drok). Myosin II interacts with actin and adherens junctions to constrict the cells on their apical sides and drive them into the embryo. In vivo studies of VF formation partly suffer from the light scattering nature of the tissue especially deep within the embryo. As a result, most of our knowledge about the functions of the genes regulating mesoderm internalization relies on antibody staining on fixed sections. Little is known about the forces and cell-biological mechanisms that dynamically regulate mesoderm internalization in vivo. Here, we report the development of an imaging-based assay to analyze ventral furrow formation in vivo. Drosophila embryos with fluorescently labeled intracellular compartments such as nucleus, 16

47th Annual Drosophila Research Conference Late Poster Abstracts membrane, junctions and cytoskeletal elements can be recorded in high spatial resolution over time by threedimensional multi-photon microscopy. The morphology of cells as well as the distribution of subcellular markers over time can be analyzed around 150 m deep within the embryo. This assay can be applied to mutant embryos and embryos, in which candidate cell-biological mechanisms have been impaired, to elucidate the molecular and cellular principles that drive mesoderm internalization in vivo. In principle, it should also be possible to adapt this assay to other morphogenetic processes in the embryo, such as the formation of the cephalic furrow or the posterior midgut. 896A Cadherin-catenin complex regulates plasma membrane localization of the exocyst complex component Sec6. Slobodan Beronja, Ulrich Tepass. Department of Zoology, University of Toronto, Toronto, ON, CANADA. The multiprotein exocyst complex, consisting of Sec3, Sec5, Sec6, Sec8, Sec10, Sec15, Exo70 and Exo84, has been shown to function in polarized exocytosis in yeast, Drosophila and vertebrate cell cultures. The exocyst acts as a tethering complex for the incoming secretory vesicles, and it is thought that by virtue of its restricted localization, the complex acts to spatially control delivery of new membrane. What controls the localization of the exocyst complex is not clear. Interactions between the apical junctional complexes and the exocyst complex have been shown in both vertebrate cell cultures and in Drosophila. The main apical junctional complex of the Drosophila epithelium is the adherens junction, components of which are the transmembrane molecule DE-cadherin (DEcad), which through homophilic interaction mediates adhesion between neighboring cells, and the cytoplasmic Dcatenin and Armadillo, which link DEcad to the actin cytoskeleton. The Drosophila genome contains two other classic cadherins: DN-cadherin (DNcad), which is largely restricted to the CNS and the non-essential DNcadherin2, which can function redundantly with DNcad. DNcad has been identified as a mediator of photoreceptor neuron targeting in the brain, and is essential for establishment of correct neuronal connections. A recent study has suggested a surprising function of the exocyst complex in the targeting process, raising the possibility that the exocyst complex functionally interacts with DNcad. In this study we are investigating the nature and functional significance of the interaction between Sec6 and the classic cadherins. 897B Endocytic control of wing development in Drosophila m. Giovanna Mottola1,2, Eric Marois1, Suzanne Eaton1, Marcos Gonzlez-Gaitn1, Marino Zerial1. 1) Max Planck Institute for Cell Biology and Genetics, Dresden, Germany; 2) Dipartimento di Biochimica e Biotecnologie Mediche, Universita' di Napoli Federico II, Napoli, Italy. Endocytosis is crucial to control cell-surface receptors and the magnitude, duration and specificity of signaling events. In the past years how endocytosis modulates signaling has been investigated mainly in cultured cells or using in-vitro cell-free systems. We want to address this question in the context of an organism such Drosophila m. We focused on the small GTPase Rab5, a key regulator of several steps of endocytosis: internalization of endocytic vesicles at the plasma membrane, fusion of these vesicles with early endosomes, early endosome biogenesis and motility. Our group has demonstrated that the multi-functionality of Rab5 is due to its ability to recruit and regulate more than 40 effectors, creating defined membrane domains. In order to follow specific Rab5-dependent endocytic steps during the development of Drosophila wing, we have explored the function of two Rab5 effectors, Rabenosyn5, already characterized in mammalian cells, and p95, one of the subunits of a novel complex. In this study we show that although part of the same machinery, Rabenosyn5 and p95 have different functions in development. Loss of Rabenosyn5 by knock out leads to lethality during first instar larval stage. Rabenosyn-5 clones are slow-growing and eliminated from the wing imaginal discs. Overexpression has also dramatic effects on the growth and patterning of the wings, which are even more severe, when apoptosis is blocked. These data highlight the relevance of Rabenosyn5 in regulating endocytosis and cell survival. On the contrary, overexpression and knock down by RNAi of p95 in Drosophila induce strong defects in wing veins formation. Analysis by immunofluorescence and confocal analysis revealed changes in the EGF dependent-activation of MAPK cascade. Other signaling pathways are not affected. Our data suggests that this novel complex has a specific involvement in the regulation of EGF receptor trafficking and/or signaling. 898C Control of Neuronal Morphogenesis and the Delta/Notch Signaling Pathway by Shrub, a Novel Coiled-Coil Protein in Drosophila. Neal T Sweeney1, 2, Jay E. Brenman3, Yuh Nung Jan2, 4, Fen-Biao Gao1, 2, 5. 1) Gladstone Institute of Neurological Disease at UCSF, San Francisco, CA; 2) Neuroscience Program, UCSF, San Francisco, CA; 3) Department of Cell and Developmental Biology and Neuroscience Center University of North Carolina,

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47th Annual Drosophila Research Conference Late Poster Abstracts Chapel Hill, NC; 4) Department of Physiology and Howard Hughes Medical Institute, UCSF, San Francisco, CA; 5) Department of Neurology, UCSF, San Francisco, CA. Dendritic branching is a complex process controlled by many signaling pathways, including Notch. Here we report the identification of mutations in the shrub gene that increased dendritic branching and enhanced signaling by the Notch pathway. Single-cell clones of shrub mutant multiple dendritic sensory neurons in Drosophila larvae showed ectopic dendritic and axonal branching in neurons, indicating a cell-autonomous function in neuronal morphogenesis. Shrub encodes an evolutionarily conserved coiled-coil protein homologous to the yeast protein Snf7, which is involved in the formation of a subset of late endosomal compartments known as multivesicular bodies. Accordingly, we found that mouse orthologs could substitute for Shrub in mutant Drosophila embryos. Loss of Shrub function caused abnormal distribution of several endosomal markers. Shrub mutant embryos also had elevated expression levels of processed Notch C-terminal fragment and a fragment of Notch ligand Delta, as well as Notch-dependent transcription. Further genetic studies revealed cell-autonomous functions for Notch and Delta in promoting dendritic branching. These data support a model in which control of dendritic morphogenesis by Shrub involves endosomal regulation of the Delta/Notch signaling pathway. 899A Domain-specific early and late function of Dpatj in Drosophila photoreceptor cells. Sang-Chul Nam, Kwang-Wook Choi. Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX 77030. The formation and maintenance of cell polarity is essential for epithelial morphogenesis. Dpatj (Drosophila homolog of mammalian Patj) is a multi-PDZ domain protein that localizes to the apical cell membrane and forms a protein complex with cell polarity proteins, Crumbs (Crb) and Stardust (Sdt). While Crb and Sdt are known to be required for the organization of adherens junctions (AJs) and rhabdomeres in differentiating photoreceptors, the in vivo function of Dpatj as a member of the Crb complex in developing eye has been unclear due to the lack of lossof-function mutations specifically affecting the dpatj gene. Our genetic analysis of hypomorph, null and RNA interference reveals distinct dual functions of Dpatj in developing and mature photoreceptors. The C-terminal region (PDZ domains 2-4) of Dpatj is not essential for development of the animal but is required to prevent late-onset photoreceptor degeneration. In contrast, the N-terminal region of Dpatj is essential for animal viability and photoreceptor morphogenesis during development. The localization and maintenance of Crb and Sdt in the apical photoreceptor membrane are strongly affected by reduced levels of Dpatj. Dpatj is necessary for proper positioning of AJs and the integrity of photoreceptors in the developing retina as well as for the maintenance of adult photoreceptors. Our study provides evidence that Dpatj has domain-specific early and late functions in regulating the localization and stability of the Crb-Sdt complex in photoreceptor cells. 900B The role of phosphatidylinositol 4-kinase III in development. Julie Tan1,2, Julie Brill1,2. 1) Developmental Biology, Hospital for Sick Children, Toronto, ON CANADA; 2) Molecular and Medical Genetics, University of Toronto, Toronto, ON CANADA. Phosphoinositides (PIPs) are involved in many fundamental cellular processes, including actin organization, membrane trafficking and cell signaling. Phosphatidylinositol 4-kinases (PI4Ks) phosphorylate the D-4 position of the inositol ring of phosphatidylinositol (PI) to yield PI4P. PI4P can be further phosphorylated to yield other PIPs such as PI(4,5)P2 and PI(3,4,5)P3. Whereas PI(4,5)P2 and PI(3,4,5)P3 are recognized as important molecules of the cell, PI4P itself is emerging as a key player in cellular trafficking, secretion and signaling. Drosophila has three PI4Ks, PI4KII, PI4KIII, and PI4KIII/Fwd. Of these, only Fwd has been characterized genetically. Fwd is required only for the completion of cytokinesis in male meiosis. fwd mutants are male sterile but viable, indicating that the other PI4Ks likely play essential roles in other tissues during development. Although PI4KIII is essential in budding yeast, where it plays roles in cell wall integrity, vacuole morphology and actin organization, the function of PI4KIII has never been studied in a multicellular organism. My long-term goal is to characterize the effect of mutations in PI4KIII on cell growth and signaling during development. Using P-element mutagenesis, I generated a deletion in the PI4KIII gene (CG10260) that removes the last 33 amino acids of the predicted catalytic kinase domain and results in lethality. I am designing rescue constructs to confirm that lethality is due to loss of PI4KIII. Also, I am using another deletion that brought the P-element closer to PI4KIII to screen for larger deletions predicted to be protein null. Future directions include dissecting the tissue specificity, molecular function and genetic interactors of PI4KIII which together will uncover the biological role of PI4KIII and PI4P in development.

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47th Annual Drosophila Research Conference Late Poster Abstracts GENOME AND CHROMOSOME STRUCTURE 901C CHD1: a chromodomain-containing chromatin remodeling factor. Jennifer A. Armstrong, Matthew S. Berger, Jennifer M. Lee, Ivy E. McDaniel, Nick L. Reeves. Joint Science Dept, Claremont Colleges, Claremont, CA. We are interested in dissecting the relative roles of the many chromatin remodeling factors localized to transcriptionally active regions. CHD1 is a member of a large family of ATP-dependent chromatin remodeling factors, including Brahma (BRM) and ISWI. CHD1 co-localizes with elongating RNA Polymerase II (Pol II) on larval salivary gland polytene chromosomes, suggesting that CHD1 may facilitate the passage of Pol II through chromatin. Human CHD1 was recently shown to bind histone H3 methylated on lysine 4 via its chromodomains, while yeast CHD1 was unable to bind methylated H3K4. The tandem chromodomains of Drosophila CHD1 are more similar to those in the human protein, suggesting that histone methylation might recruit CHD1 to active genes in Drosophila. To understand interactions between CHD1 and other chromatin remodeling factors, we are making use of eye-based assays designed to identify factors important for the BRM and ISWI chromatin remodeling factors. Preliminary studies indicate that loss of brm makes cells more sensitive to levels of chd1. In addition, CHD1 localization on polytene chromosomes suggests that its activity may not be limited to Pol II genes. We find moderate levels of CHD1 in the nucleolus during normal larval growth, and high levels following heat shock, suggesting that CHD1 may be involved in transcription of rRNA genes. 902A Deciphering chromatin based cellular processes in development using a dominant negative histone acetyltransferase. Meridith Toth, Xianmin Zhu, Neetu Singh, Felice Elefant. Dept Bioscience/Biotechnology, Drexel Univ, Philadelphia, PA. DNA is packaged into nucleosomes around histone protein cores so tightly that post-transcriptional modifications are necessary to allow gene expression to proceed. Targeted gene expression which is critical in differentiation and development is made possible by one such modification, acetylation, which is associated with active gene domains. Acetylation is catalyzed by histone acetyltransferases (HATs), proteins that transfer the acetyl group from acetyl coA to conserved lysine residues on histone protein tails. Misregulation of HATs early in development has implicated them in various neurodegenerative diseases and cancers. The goal of this study is to understand the specific developmental implications that a particular HAT protein plays in eukaryotic development using a Drosophila model system. Our HAT of interest is Drosophila HAT Tip60 (dmTip60), a specific human HAT homologue found by our lab through database searches. Alignments with the human homologue showed dmTIP60 to be 56%identical/68%similar, making it a good candidate for ultimately relating our findings to the human system. We have chosen this HAT because it is essential in cell cycle progression, is implicated in disease states including Alzheimers Disease and various cancers, has been directly associated with prostate cancer, but has not yet been studied in a multicellular developmental model system. We will be considering the developmental significance of this protein through the generation of dominant negative mutations, which will be expressed in vivo using a GAL4/UAS inducible system in Drosophila. The use of dominant negative mutations will allow us to differentiate between functions that are complex related and those that are based on the HAT activity of Tip60. We expect that tissue-specific production of these proteins during Drosophila embryogenesis will lead to severe phenotypic changes, and in future experiments the genes associated with these changes can be determined. 903B A dominant negative form of the BEAF insulator proteins disrupts chromatin structure. Craig M. Hart, Matthew Gilbert, Swarnava Roy, Yian Yee Tan. Dept Biological Sciences, Louisiana State Univ, Baton Rouge, LA. The division of chromosomes into discrete functional domains in the three dimensional space of the nucleus is thought to play an important role in the regulation of gene expression. The boundaries of domains are thought to actively defined by specialized DNA sequences known as insulator (or boundary) elements. In transgene assays, insulators prevent interactions between enhancers and promoters when inserted between them. Insulators also buffer bracketed transgenes against chromosomal position effects. Binding sites for the Boundary ElementAssociated Factor BEAF are necessary for the insulator activity of the scs insulator in transgenic flies. BEAF binds to hundreds of interbands on polytene chromosomes, indicating that BEAF-utilizing insulators are common in Drosophila. BEAF is comprised of complexes of two 32 kDa proteins (BEAF-32A and 32B) that are derived from the same gene, and differ only in their DNA binding domains. 32A and 32B interact with each other via a carboxyterminal domain, but do not form stable complexes with other proteins. We have made transgenic flies with a transgene encoding the BEAF self-interaction domain (BID) which is under GAL4 UAS control. We provide

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47th Annual Drosophila Research Conference Late Poster Abstracts evidence that the BID protein functions as a dominant negative form of BEAF. In support of models that propose a relationship between insulator function and chromatin, results will be presented that show that BID affects chromatin structure or dynamics. 904C Identification and Characterization of Gypsy-like Endogenous Insulators in Drosophila melanogaster. Edward Ramos, Kelly Baxter, Victor Corces. Dept of Biology, Johns Hopkins University, Baltimore, MD. Chromatin insulators have been implicated in the regulation of higher-order chromatin structure and may function to compartmentalize the eukaryotic genome into independent domains of gene expression. To test this possibility, we used biochemical and computational approaches to identify gypsy-like genomic binding sites for the Suppressor of Hairy-wing [Su(Hw)] protein, a component of the gypsy insulator. The gypsy insulator has long been known to prevent enhancer promoter communication but most studies done use the insulator initially found in the gypsy retrotransposon rather than endogenous binding sites. To increase our knowledge of insulators in the Drosophila genome, we set out to characterize endogenous gypsy-like Su(Hw) binding sites. EMSA and FISH analyses suggest that these newly identified gypsy-like insulators are genuine Su(Hw) binding sites and thus function as bona fide insulators. The insulator strength is dependent on the genomic location of the transgene and the number of Su(Hw) binding sites, with clusters of 2-3 sites showing a stronger effect than individual sites. These clusters of Su(Hw) binding sites are mostly located in intergenic regions or in introns of large genes, an arrangement that fits well with their proposed role in the formation of chromatin domains. Taken together, these data suggest that genomic gypsy-like insulators are capable of regulating gene expression and may provide a means for the compartmentalization of the genome within the nucleus. 905A Mapping and mutagenesis of the E(var)3-9 locus. Karen Weiler. Dept of Biology, West Virginia University, Morgantown, WV. E(var)3-9 is a member of a large class of genetic modifiers of PEV identified by mutations that dominantly enhance the variegation of wm4. The mutant phenotype suggests a role for the wild-type E(var)3-9 protein in antagonizing heterochromatin formation. E(var)3-9 is also required for female fertility. Initial meiotic recombination and deficiency mapping localized E(var)3-9 to either of two intervals, 85C1-2 to 85D8 or 85F6 to 86C1. The results of P element-mediated male recombination mapping confirmed that E(var)3-9 resided within 85C-D but were unable to narrow the target significantly, in part due to ambiguous data. However, the availability of new deficiencies synthesized using P element insertions facilitated the localization of E(var)3-9 to a region encompassing twenty-three known and predicted genes. The results of additional genetic approaches taken to identify the E(var)3-9 gene will be described. These include using the FLP-FRT deletion generation system (1) and the P{wHy} deletion-generator system (2) to create novel deficiencies. E(var)3-9 was determined to be allelic to CG11971, which is predicted to encode a zinc-finger protein. 1) Parks et al. (2004) Nat Genet 36(3):288 2) Huet et al. (2002) PNAS 99(15):9948. 906B Transmission of Fragment Chromosomes Lacking An Endogenous Telomere. Simon Titen, Kent Golic. Dept Biol, Univ Utah, Salt Lake City, UT. It was first shown by Herman Muller and Barbara McClintock, in the 1930s and 40s, that the ends of linear chromosomes have distinct properties. An end, or a telomere, is comprised of both protein and DNA that accomplish two specific functions: 1) Prevent the cell from recognizing the end of the chromosome as a DNA double strand break (DSB), which subsequently prevents end-to-end fusions, cell cycle arrest, and interactions between new DSBs and chromosome ends; and 2) Solve the end replication problem. In order to characterize the mechanisms involved in telomere generation and maintenance our lab has developed a technique to generate chromosomes that lack an endogenous telomere, in vivo, by inducing dicentric bridge formation using the site-specific recombinase FLP. We have shown that, in the male germ line, these bridges frequently break and the two resulting fragments are transmitted to the progeny. Our research has been focused on characterizing the molecular structure of these fragment chromosomes and the proteins required for de novo telomere addition and germ line transmission. 907C Characterizing Drosophila telomeres that lack natural transposons. Xiao-Feng Zheng, Yikang Rong. Lab of Molecular Cell Biology, National Cancer Institute, Bethesda, MD. Drosophila telomeres are maintained by targeted attachments of retrotranspons to chromosomal ends, as oppose to telomere elongation by telomerase in most eukaryotes. It is of great interest therefore, to characterize the 20

47th Annual Drosophila Research Conference Late Poster Abstracts structure and function of Drosophila telomeres, drawing potential similarities to those of popular telomeres. Characterizing Drosophilas natural telomere is unfeasible however, due to sequence heterogeneity at the ends. To circumvent this limitation, we created a telomere with a defined sequence by generating a double strand break to eliminate the telomere-associated sequence and retrotransposons. Stocks with this terminally-deleted chromosome can be stably maintained, even with continuous degradation of DNA at the end. The new ends are sometimes healed by attachments of transposons. We have initiated structural analysis of this unique end. Preliminary results suggest that Drosophila telomeres can also exist in a terminally single-stranded conformation. 908A Partially redundant roles of Drosophila ATM and ATR checkpoint kinase for telomere maintenance. Xiaolin Bi1, Deepa Srikanta1, Laura Fanti2, Sergio Pimpinelli2, Yikang Rong1. 1) NCI/NIH, Bethesda, MD,USA; 2) Dipartimento di Genetica e Biologia Molecolare,Universita di Roma"La Sapienza",Rome,Italy. In higher eukaryotes, the ataxia telangiectasia mutated (ATM) and ATM and Rad3-related (ATR) checkpoint kinases play distinct, but partially overlapping, roles in DNA damage response. Yet their interrelated function has not been defined for telomere maintenance. We discover in Drosophila that the two proteins control partially redundant pathways for telomere protection: the loss of ATM leads to the fusion of some telomeres, whereas the loss of both ATM and ATR renders all telomeres susceptible to fusion. The ATM-controlled pathway includes the Mre11 and Nijmegen breakage syndrome complex but not the Chk2 kinase, whereas the ATR-regulated pathway includes its partner ATR-interacting protein but not the Chk1 kinase. This finding suggests that ATM and ATR regulate different molecular events at the telomeres compared with the sites of DNA damage. This compensatory relationship between ATM and ATR is remarkably similar to that observed in yeast despite the fact that the biochemistry of telomere elongation is completely different in the two model systems. We provide evidence suggesting that both the loading of telomere capping proteins and normal telomeric silencing requires ATM and ATR in Drosophila and propose that ATM and ATR protect telomere integrity by safeguarding chromatin architecture that favors the loading of telomere-elongating, capping, and silencing proteins.

REGULATION OF GENE EXPRESSION 909B A mutational analysis of ecdysone receptor and Ultraspiracle interaction in a heterologous cell culture system. Vincent C Henrich, Josh Beatty, Jenna Callender. Inst. Health, Science, and Soc, UNC-Greensboro, Greensboro, NC. The ecdysone receptor and the insect RXR homologue, Ultraspiracle (USP) form a heterodimer that mediates transcriptional activity in the absence and presence of the insect steroid hormone, 20-hydroxyecdysone. Plasmids encoding these receptors were transfected and expressed in a mammalian cell line that normally shows no response to ecdysteroids or juvenile hormone, so that an ecdysteroid response system was reconstituted for analysis. Several site-directed mutations have been produced for each of the three natural EcR isoforms and two constructs of USP, one encoding the DNA-binding domain and the other without it, by measuring their effects on basal, ecdysteroid-induced, and juvenile hormone potentiated transcriptional activity in cells. The sites were selected based on known or suspected effects on transcriptional activation functions, cofactor interactions, dimerization, and ligand-binding, using sequence alignments, computational models, and previous studies. The studies revealed that some EcR mutations impair specific receptor subfunctions whereas most USP mutations tend to either obliterate activity or exert little effect at all. The analysis indicates that various aspects of EcR/USP transcriptional activity via the canonical hsp27 ecdysone response element depend upon EcR isoform, the presence or absence of the USP DNA-binding domain, and specific residues within the USP ligand-binding domain. 910C Disruption of gene function by targeted insertion and inducible RNAi demonstrate nautilus is essential for embryonic myogenesis and viability and impacts founder cell patterning. Qin Wei1, Yikang Rong2, Bruce Patterson1. 1) Laboratory of Biochemistry, NCI/NIH, Bethesda, MD; 2) Laboratory of Cellular and Molecular Biology, NCI/NIH, Bethesda, MD. The nautilus gene is the single member of the bHLH family of myogenic regulatory factors in Drosophila. Embryos injected transiently with nautilus dsRNA displayed severe embryonic muscle disruption indicating nautilus is essential for embryonic muscle formation. In order study nautilus loss-of function throughout development, we used two complementary approaches to inactivate gene function: in the first instance, we used targeted gene disruption to introduce an armadillo-GFP expression cassette in the middle of the first exon of nautilus gene; in the second, we utilized an inducilbe RNAi transgene to produce tissue-specific expression of nautilus dsRNA under the control of gal 4. Both the targeted nautilus null and the RNAi transgene produce the same range of muscle defects in the 21

47th Annual Drosophila Research Conference Late Poster Abstracts embryo, from the severe loss of muscle organization, similar to the RNAi phenotypes observed with the injection of dsRNA, to minor muscle pattern disruptions. All stages of development are affected: larvae are weak and move slowly in an uncoordinated fashion, the majority of pupae are unable to exit the pupal case and the small precentage that do usually die shortly after hatching. Less than 5% of the homozygous embryos survive to adults and females are sterile with enlarged abdoments unable to lay eggs. An hsp70 nau-cDNA transgene rescues the null. nautilus precedes duf expression in founder cells at stage 10 but they are co-exprressed by stage 13. The embryonic muscle disruption correlates with an abnormal founder cell pattern. These results demonstrate conclusively that the nautilus gene is required for normal embryonic myogenesis and viability but is not required for the determination of the myogenic cell lineage. 911A

Eif1A mutations dominantly suppress the eggshell patterning defect caused by activation of the meiotic recombination checkpoint. Martha Klovstad1, Gail Barcelo1, Uri Abdu2, Trudi Schpbach1. 1) HHMI, Molecular Biology Dept., Princeton University, Princeton, NJ; 2) Dept. of Life Sciences, Ben-Gurion University. During early Drosophila oogenesis double-stranded breaks (DSBs) are formed as an early intermediate in meiotic recombination. Analysis of the spindle class mutants has shown that the persistence of unrepaired DSBs during Drosophila oogenesis leads to activation of a meiotic recombination checkpoint. Similar checkpoints act in yeast, C.elegans, and mice and result in the induction of apoptosis in mice and C.elegans and in cell cycle arrest in yeast. In contrast, checkpoint activation in Drosophila results in the ventralization of the eggshell and oocyte nuclear morphology defects. Specifically, checkpoint activation results in the inhibitory modification of the translation factor Vasa, a protein which is required for translation of the DV patterning ligand Gurken. In order to identify genes involved in the meiotic recombination checkpoint we have undertaken an EMS mutagenesis screen on the third chromosome for dominant and recessive genetic modifiers of the eggshell defect of mutants in spindleB, a spindle class gene required for DSB repair. One gene identified as a dominant suppressor is the translation factor, eif1A. Double mutants for spnB and eif1A lay predominately wild-type like eggs. Gurken antibody staining shows that localized Grk protein levels are restored in the majority of double-mutant egg chambers. Eif1A has been implicated in 80S ribosome assembly, ribosome scanning, and stabilizing the initiator Met-tRNA on the 40S ribosomal subunit. Most notably, Eif1A binds Eif5B in other species and Eif5B has previously been shown to regulate Vasa function. We are currently determining the specificity of the eif1A/spnB interaction and examining potential functional interactions between Eif1A, Eif5B and Vasa .
912B Regulation of Histone Acetyltransferase during Development. Vijayalakshmi Ramakrishnan. Drexel University, Philadelphia, PA. Histone Acetyltransferases (HATs) are a key class of chromatin regulatory enzymes that acetylate the positively charged lysine tails of nucleosomal histones. This process serves to decondense higher order chromatin structure which in turn, activates gene expression. Although it has been established that the misregulation of HATs results in deleterious developmental defects, the mechanism underlying their regulation during development remains unclear. We have previously demonstrated that two human HAT homologs in Drosophila, dmELP3 and dmTIP60, are differentially expressed during Drosophila development, suggesting that HAT activity is controlled, at least in part, by regulation of their expression during development. To extend these studies, we are currently using an in situ hybridization technique to delineate the tissue and cell type specific expression patterns of these HATs. Our preliminary results demonstrate that both dmELP3 and dmTIP60 HATs are also differentially expressed in specific tissues and cell types during various stages of development, suggesting that HAT activity is also controlled at the level of tissue and cell-type specific HAT expression profiles. 913C Functional characterization of the histone acetyltransferase Elp3 during development. Neetu Singh, Felice Elefant. Dept Bioscience/Biotechnology, Drexel University, Philadelphia, PA. Histone acetyltransferases (HATs) are a class of enzymes that modify chromatin packaging to facilitate gene expression; however their role during development remains unclear. We are focusing our studies on the Drosophila HAT (dmHAT) homolog Elp3, as it demonstrates high homology to human Elp3, which plays an essential role in transcriptional activation and elongation, and contains a putative demethylase domain. We first identified dmElp3 through database searches and subsequently cloned the cDNA encoding this gene. Alignments between the dmElp3 and human Elp3 proteins demonstrate significant homology: 82% identity, 92% similarity, supporting the use of this homolog in deciphering human Elp3 developmental function. To characterize dmElp3 function, we are using an inducible GAL4/RNAi based system in Drosophila to silence endogenous dmELP3 expression in a variety of tissues and developmental stages. A 650bp non-conserved fragment from dmELP3 was cloned into a senseantisense inverted gene arrangement under the control of UAS binding sites to induce RNA knockdown of 22

47th Annual Drosophila Research Conference Late Poster Abstracts endogenous dmELP3. A control construct was created in which the inverted repeat fragments were cloned in a sense-sense orientation. Transgenic flies carrying the ELP3/RNAi construct were created to induce RNAi expression in a ubiquitous and tissue specific manner. Our preliminary results demonstrate that ubiquitous expression of ELP3 RNAi produces a slow growth phenotype and embryonic lethality, suggesting Elp3 is required for proper development. We are currently creating transgenic flies carrying the entire dmELP3 ORF under the control of UAS binding sites to investigate the effects of inducing dmElp3 overproduction in specific tissues during development. Assays will be performed to determine levels of histone acetylation and demethylation after altering endogenous dmElp3 production. Aberrant phenotypes resulting from altering production of dmElp3 will be correlated to known developmental pathways to aid in deciphering the cellular function of dmElp3 in chromatin control during development. 914A Functional analysis of the chromatin regulator protein: TIP60. Xianmin Zhu, Felice Elefant. Dept Bioscience/Biotechnology, Drexel Univ, Philadelphia, PA. Histone Acetyltransferases (HATs) are a key class of chromatin regulatory proteins that mediate developmental chromatin control, however little is known about their roles during development. We are focusing our studies on the HAT TIP60, as it has been shown to play essential roles in transcription and cell cycle control. We have identified and cloned the human HAT homolog of TIP60 in Drosophila which we are using to decipher human HAT developmental function in the fly model system. Alignment of dmTip60 with the human Tip60 protein demonstrates significant homology: dmTIP60 is 56% identical/ 68% similar. To investigate the mechanism(s) underlying developmental HAT regulation, we have examined the developmental expression profiles of dmTIP60 using a realtime RT-PCR assay. These studies reveal that dmTip60 is differentially expressed during Drosophila development, with levels peaking during embryogenesis. Our results suggest that dmTIP60 HAT activity is controlled, at least in part, through their developmental regulation, and indicate the importance of such activity in early development. To decipher the roles of such dmHATs during embryogenesis, we are using a GAL4 targeted RNAi based system in Drosophila to silence endogenous dmTip60 expression in a variety of tissues and developmental stages of choice. Transient transfection data demonstrates that our TIP60/RNAi HAT knockdown construct results in deleterious effects in the embryonic Drosophila S2 cell line, suggesting that Tip60 is required during embryogenesis. We have generated three independent transgenic fly lines for each of our TIP60/RNAi and control constructs and induced their expression using an actin GAL4 driver fly line (Act5-GAL4). Our results show that actin-specific downregulation of TIP60 results in lethality of all progeny, supporting an essential role for Tip60 in chromatin control during development. 915B Functional analysis of promoter-enhancer interactions at the Drosophila bithorax complex. Omar S Akbari, Esther Bae, Robert Drewell. Biology, University of Nevada Reno, Reno, NV. The Drosophila bithorax complex (BX-C) consists of over 300 kb of DNA, but encodes only 3 homeotic genes. The extensive intergenic infrabdominal (iab) region is known to harbor cis-regulatory elements capable of directing the segment-specific expression patterns of the homeotic genes. We have been examining the mechanisms which regulate promoter-enhancer interactions at the BX-C. There are many examples within gene complexes of shared transcriptional enhancers interacting only with a subset of target promoters. Promoter competition and insulators are thought to play a role in regulating these interactions. In the bithorax complex of Drosophila, the IAB5 enhancer is located 55 kb 3 of the Abdominal-B promoter and 48 kb 5 of the abdominal-A promoter. Although roughly equidistant from the two promoters, IAB5 specifically interacts only with the Abdominal-B promoter, even though the enhancer and promoter are separated by at least two insulators. Our experiments demonstrate that a novel 254bp cis-regulatory element located 5 of the Abd-B transcriptional start site is able to tether IAB5 to the Abd-B promoter in transgenic embryo assays. Furthermore, the promoter tethering element (PTE) sequence is sufficient to direct IAB5 to an ectopic site in competition assays. These results provide in vivo evidence that specific long-range enhancer-promoter interactions in the bithorax complex are regulated by a PTE located 5 of the Abd-B promoter and suggest that tethering elements might be a common epigenetic mechanism for the regulation of enhancerpromoter specificity at other gene complexes.

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47th Annual Drosophila Research Conference Late Poster Abstracts SIGNAL TRANSDUCTION 916C Drosophila TAB2 is required for the immune activation of JNK and NF-kappaB. Baoxue Ge1, Zi-Heng Zhuang1, Lei Sun1, Ling Kong1, Jun-Hao Hu1, Ming-Can Yu1, Peter Reinach2, Jin-Wu Zang1. 1) Institute of Health Sciences, Shanghai, CHINA; 2) Department of Biological Sciences, SUNY College of Optometry, New York, NY 10036, USA. The TAK1 plays a pivotal role in the innate immune response of Drosophila by controlling the activation of JNK and NF-kappaB. Activation of TAK1 in mammals is mediated by two TAK1-binding proteins, TAB1 and TAB2, but the role of the TAB proteins in the immune response of Drosophila has not yet been established. Here, we report the identification of a TAB2-like protein in Drosophila called dTAB2. dTAB2 can interact with dTAK1, and stimulate the activation of the JNK and NF-kB signaling pathway. Furthermore, we have found that silencing of dTAB2 expression by dsRNAi inhibits JNK activation by peptidoglycans (PGN), but not by NaCl or sorbitol. In addition, suppression of dTAB2 blocked PGN-induced expression of antibacterial peptide genes, a function normally mediated by the activation of NF-kappaB signaling pathway. No significant effect on p38 activation by dTAB2 was found. These results suggest that dTAB2 is specifically required for PGN-induced activation of JNK and NF-kappaB signaling pathways. 917A Regulation of Drosophila p38 activation by specific MAP2 kinase and MAP3 kinase in response to different stimuli. Baoxue Ge1, Zi-Heng Zhuang1, Yuan Zhou1, Ming-Cai Yu1, Neal Silverman2. 1) Institute of Health Sciences, Shanghai, CHINA; 2) Division of Infectious Disease, Department of Medicine, University of Massachusetts Medical School, 364 Plantation St, Worcester MA 01605, USA. The p38 mitogen-activated protein kinase (MAPK) signaling pathway plays an important role in cellular responses to inflammatory stimuli and environmental stress. Activation of p38 is mediated through phosphorylation by upstream MAPKK, which in turn is activated by MAPKKK. However, the mechanism of how different upstream MAP2Ks and MAP3Ks specifically contribute to p38 activation in response to different stimuli is still not clearly understood. By using double-stranded RNA-mediated interference (RNAi) in Drosophila cells, we demonstrate that D-MKK3 is a major MAP2K responsible for D-p38 activation by UV, heat shock, NaCl or peptiodglycan (PGN). Stimulation of UV and PGN activates D-p38 through D-MEKK1, heat shock-induced activation of D-p38 signals through both D-MEKK1 and D-ASK1. On the other hand, maximal activation of D-p38 by NaCl requires the expression of four MAP3Ks. 918B Role of PP2A in visual signaling. Ning Wang, Bih-Hwa Shieh. Dept of Pharmacology, Vanderbilt University, Nashville, TN. Visual signaling is a rhodopsin-mediated event that leads to depolarization of photoreceptors by opening TRP and TRPL channels. Specifically, rhodopsins activate Gq that turns on a phospholipase C (NORPA) to catalyze the breakdown of PIP2 to generate IP3 and DAG. DAG is a potent activator of eye-PKC that modulates the visual response. Eye-PKC is tethered to a macromolecular complex that contains the scaffolding protein INAD, NORPA and TRP. Previously, eye-PKC has been shown to phosphorylate both TRP and INAD in vitro. Phosphorylation of TRP and INAD is likely to contribute to the negative regulation of the visual response as mutants lacking eye-PKC display slow deactivation and abnormal desensitization. To counteract the effects of eye-PKC, we investigated the role of protein phosphatases in dephosphorylation of INAD and TRP by biochemical analysis. We show that dephosphorylation of INAD was suppressed by PP2A/PP1 inhibitor okadaic acid. To isolate the critical protein phosphatases for INAD, we employed FPLC and found that the C subunit of PP2A (PP2Ac) was co-purified with the phosphatase activity of INAD. Consistently, recombinant human PP2Ac is able to dephosphorylate INAD in vitro,whereas PP1c cannot. By immunoprecipitation, PP2Ac is also co-purified with the INAD complex and can be pull-downed by INAD. We examined thein vivofunctions of PP2A by characterizing hypomorphic alleles of PP2Ac using electroretinograms. We show that an accelerated deactivation of the visual response in flies expressing a lower amount of PP2Ac. These results indicate that PP2A modulates the visual response probably by counteracting the effects of eye-PKC on INAD and TRP. 919C Drosophila glypican Dally-like is essential for Breathless mediated tracheal development in embryos and wing discs. Dong Yan1,2, Xinhua Lin1,2. 1) Division of Developmental Biology, Cincinnati Children's Hospital, Cincinnati, OH; 2) Molecular and Developmental Biology Graduate Program, University of Cincinnati, OH. Genetic studies in Drosophila have shown that heparan sulfate proteoglycans (HSPGs) are involved in both breathless (btl) and heartless (htl) mediated FGF signalling during embryonic development. But it is not known which HSPG core protein is involved in these events. We have demonstrated that dally-like (dlp, one Drosophila 24

47th Annual Drosophila Research Conference Late Poster Abstracts glypican) mutant embryos have tracheal migration defects similar to mutants of btl. Moreover, btl-dependent MARK activation is significantly reduced in dlp mutant embryos. However, htl-dependent FGF signalling and mesodermal migration are not affected in dlp mutant embryos. Ectopic expression of other proteoglycan core proteins, such as Dally, Syndecan and Perlican, fail to rescue tracheal phenotype in dlp mutant embryos. Together, these new results suggest Dlp is specific for btl-mediated FGF signalling in embryo. We further analyze the role of Dlp in air sac tracheoblast cell migration in wing disc, another event mediated by Bnl/ Btl signalling. To our surprise, we find HSPGs are dispensable in Bnl expression cells, while play essential role exclusively in Bnl receiving cells, the air sac tracheoblast cells. Dlp has a redundant function with another glypican, Dally, to receive FGF signalling in air sac cells. These novel results provide clear evidence that HSPG is only required in FGF receiving cells rather than secretion cells in vivo, an issue that is difficult to address by biochemical studies and other model systems. Our mechanistic data favour a model in which HSPGs facilitate FGF/FGFR interaction and/or stabilization. 920A The Rac1 and Rac2 C-terminal Regions Interact Differently with dPosh and Ter94. Li Xiao, Douglas Ruden. Environmental Health Sciences, University of Alabama at Birmingham, Birmingham, AL. Rac1 and Rac2 are closely-related small G proteins that participate in numerous signaling pathways, including JNK activation. The RING domain-containing Posh (Plenty of SH3 domains) is a scaffold protein that interacts with components of the JNK and Akt signaling pathways. Vertebrate Posh binds Rac1 through its putative Rac1 binding domain (RBD), but this domain does not exist in Drosophila Posh (dPosh). Ter94 is a Drosophila homolog of VCP, which is a molecular chaperon involved in a variety of cellular functions. We found that mutations of Rac1, Rac2, dPosh, and Ter94 affect Drosophila rhabdomere structure, suggesting that theses proteins are involved in eye development. Eye morphology studies indicate that both Rac1 and Rac2 genetically interact with dPosh. Interestingly, either dPosh over-expression or decreased-expression enhances the Rac1 and Rac2 over-expression eye-degeneration phenotypes, suggestting that a proper dose of dPosh is required for balancing apoptosis and survival signals. Overexpressing RING-mutant dPosh almost totally loses its enhancer role with overexpressed Rac2, but it still enhances the eye-degeneration phenotype of overexpressed Rac1. The main difference of Drosophila Rac1 and Rac2 is the C-terminal polybasic region which is important for their subcellular localization. If the polybasic regions of Rac1 and Rac2 are swapped, overexpressing wild type dPosh still enhances the chimeric Rac overexpression phenotypes. However, over-expressing the RING-mutant-dPosh does not enhance of chimeric Rac2/1 (Rac2 with the C-terminal tail of Rac1), suggesting that the C-terminal polybasic region of Rac2 is sufficient for dPosh RING-mediated specific interactions. Genetic studies also show that Ter94 specifically interacts with Rac2, but not Rac1, and the Rac2 polybasic region is required for the specific Ter94-Rac2 interaction. These results might be partly explained because Rac2 and Ter94, and possibly dPosh, are predominantly membrane associated proteins, whereas, Rac1 is mostly cytosolic. 921B An investigation of the oligomerization properties of SNS during myoblast fusion. Kiranmai Kocherlakota1,2, Erika Geisbrecht1, Susan Abmayr1. 1) Stowers Institute for Medical Research, 1000 E 50th Street, Kansas City, MO 64110; 2) Cell and Developmental Biology Department, Huck Institute of Life Sciences, Pennsylvania State University, PA 16802. The cell adhesion molecule Sticks-and-Stones (SNS) serves as a receptor on the surface of fusion competent myoblasts during the process of myoblast fusion in Drosophila embryos. SNS is absolutely essential for myoblast fusion and mutants are characterized by complete absence of mature muscle fibers. Several cell adhesion molecules use oligomer formation as a mechanism to enrich into discrete punctae for strong adhesive contacts. Such protein redistribution has indeed been observed for SNS. In transiently transfected Schneider line 2 (S2) cells, SNS is distributed uniformly throughout the plasma membrane but redistributes and localizes to points of cell contact when aggregated with cells expressing its ligands. On the surface of myoblasts in the embryonic mesoderm, SNS is present in discrete punctae that colocalize with its ligands (Galletta et al. Mech Dev 2004, 121:1455-68). In an effort to understand SNS function during myoblast adhesion and signaling, we have investigated its oligomerization properties. SNS forms homo-oligomers in S2 cells and in the embryonic mesoderm. Deletion of the cytodomain and the extracellular domain of SNS did not affect oligomer formation of SNS. Oligomerization requires only the transmembrane sequences. In addition, evidence for SNS hetero-oligomerization with its paralog Hibris (Hbs) came from mass spectrometric analysis of affinity purified SNS. This interaction was

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47th Annual Drosophila Research Conference Late Poster Abstracts confirmed in the embryonic mesoderm by co-immunoprecipation and western blotting. Hibris (Hbs) is expressed on the surface of at least a subset of fusion competent myoblasts. One of the biological implications of SNS oligomerization may be association into membrane microdomains such as lipid rafts to enrich SNS for strong adhesive contacts. Currently we are investigating this possibility by biochemical fractionation and fluorescent labeling techniques. 922C The Ihog cell surface proteins bind and mediate response to the Hedgehog signal. Xiaoyan Zheng2, Shenqin Yao2, Lawrence Lum2, 4, Philip Beachy1, 2, 3. 1) Howard Hughes Medical Institute; 2) Molecular Biology and Genetics, The Johns Hopkins University School of Medicine, Baltimore, MD; 3) Department of Oncology, The Johns Hopkins University School of Medicine, Baltimore, MD; 4) Current address: Department of Cell Biology, Univeristy of Texas Southwestern Medical Center, Dallas, TX. The ihog gene (interference hedgehog), identified by RNA interference in Drosophila cultured cells, encodes a type 1 membrane protein shown here to bind and mediate Hedgehog (Hh) signal response. ihog mutations produce defects characteristic of Hh signaling loss in embryos and imaginal discs, and epistasis analysis places ihog action at or upstream of the negatively-acting receptor component, Patched (Ptc). The first of two extracellular fibronectin type III (FNIII) domains of the Ihog protein mediates a specific interaction with Hh protein in vitro, but the second FNIII domain is additionally required for in vivo signaling activity and for Ihog-enhanced binding of Hh protein to cells co-expressing Ptc. Other members of the Ihog family, including Drosophila Boi and mammalian CDO and BOC, also interact with Hh ligands via a specific FNIII domain, thus identifying an evolutionarily conserved family of membrane proteins that function in Hh signal response. 923A Reconstitution of Hedgehog signaling from exogenously introduced pathway components. Sekyung Oh, Philip A. Beachy. Molecular Biology & Genetics, Howard Hughes Medical Institute, The Johns Hopkins University School of Medicine, Baltimore, MD. Genetic studies have identified Hedgehog (Hh) signaling components and assigned their pathway roles. To better understand the mechanisms of transduction in cells responding to the Hh signal, we reconstituted pathway activity in otherwise unresponsive S2R+ cultured cells by expression of exogenously introduced pathway components. S2R+ cells are unresponsive because they lack the zinc finger transcription factor Cubitus interruptus (Ci), the major transcriptional effector of the pathway. However, transfection of Ci alone does not restore Hh-regulated response because cellular factors required for its proteolytic processing, intracellular localization, and/or transcriptional activation are insufficiently expressed. Co-transfection of the four core pathway components Smoothened (Smo), Costal2 (Cos2), Fused (Fu), and Suppressor of fused (Su(fu)) not only restores Hh-dependent Ci regulation but also results in the expected posttranslational modifications of all individual components. This system will be useful for investigating mechanisms of Hh signal transduction because of its dependence on exogenously introduced components, which can be altered in vitro to examine specific aspects of their function. 924B Drosophila cysteine peptidase Tan: expression in photoreceptor cells R1-R8 lends support for a putative histamine neurotransmitter / carcinine cycle between photoreceptor terminals and epithelial glia. Bernhard Hovemann1, Stefanie Wagner1, Christiane Heseding1, John True2. 1) Dept Chemistry, Ruhr Univ, Bochum, Bochum, GERMANY; 2) Department of Ecology and Evolution, State University of New York, Stony Brook, NY 11794-5245, USA. The Drosophila mutant tan (t) shows reciprocal pigmentation defects when compared to ebony (e). Visual phenotypes, however, are related in both mutants: Electroretinogram (ERG) recordings lag on and off transients, an indication of impaired synaptic transmission to postsynaptic cells L1 and L2. Recent cloning of tan revealed transcription of the gene in the retina, most likely in photoreceptor cells. We expressed Tan in E.coli and confirmed by Western blotting and mass spectroscopic analysis that Tan like its fungal ortholog isopenicillin-N Nacyltransferase is expressed as preprotein followed by autoproteolytic cleavage into two subunits at a conserved Gly-Cys motif. This assigns Tan to the family of Cysteine Peptidases. To disclose expression of Tan as opposed to Ebony in the optic lobe antisera against Tan peptides were raised. Co-localization with glial and neuronal cell markers revealed neuronal expression of Tan that is confined to the cytosolic fraction of all photoreceptor cells. Strong immunoreactivity extends to the axonal termini in lamina and medulla. A close proximity of Tan and Ebony expression is particularly evident in lamina cartridges where three epithelial glia cells envelop the six photoreceptor termini R1-6. This local proximity lends support for a model in which Tan and Ebony interact biochemically in a putative histamine inactivation and recycling pathway in Drosophila vision.

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47th Annual Drosophila Research Conference Late Poster Abstracts PATTERN FORMATION 925C Identification of a novel gene that is regulated by engrailed and functions in Drosophila embryo segmentation. Hyunsuk Suh, Jeongbin Yim. Biological Sciences, Seoul National University, Seoul, KOREA. Pattern formation in Drosophila embryo is proceeded by sequential actions of maternal, gap, pair-rule, and segment polarity genes (i.e. wingless, engrailed) and eventually give rise to repeating segmental stripes that subdivide the anterior-posterior axis. In order to search for novel genes that are expressed in specific patterns, enhancer trap screening was carried out and a candidate gene that is expressed in a segment-specific manner was obtained. Expression pattern during each embryonic stage was analyzed by RNA in situ hybridization. Interestingly, it is expressed in a series of wide band of cells that overlap parasegment boundaries during stage 7-10, while in later stages, it is only expressed in cells neighboring engrailed (en)-expressing cells. Expression was also downregulated in cells ectopically expressing en in paired-Gal4>UAS-en embryos. Denticle belt fusion was observed (penetrance:~5%) in embryos injected with various concentrations of dsRNA which target unique regions in the open reading frame of the candidate gene. In summary, we have identified a novel gene that is negatively regulated at transcription level by engrailed and is required for proper segmentation in Drosophila embryo. 926A Kinesin and Dynein are required for anterior-specific bicoid mRNA transport in the Drosophila oocyte. Byeong J. Cha1, William E. Theurkauf2. 1) Dept Cell & Developmental Biol, Vanderbilt Univ Medical Ctr, Nashville, TN; 2) Program in Molecular Medicine, University of Massachusetts Medical School, Worceter, MA. bicoid mRNA is synthesized in the nurse cells and is transferred to the oocyte, where it localizes to the anterior cortex. bicoid mRNA injected into the nurse cells and transferred to the oocyte localize specifically to the anterior cortex, while bicoid mRNA injected directly into the oocyte shows non-polar movement to the lateral and anterior cortex. Mutations in dynein heavy chain or anti-dynein antibodies block both anterior-specific movement of bicoid mRNA complexes formed in the nurse cells and non-polar movement of transcript injected directly into the oocyte. A null kinesin heavy chain mutation has no effect on non-polar transport of bicoid mRNA injected directly into the oocyte, and bicoid mRNA injected into a kinesin heavy chain mutant nurse cell and transferred to a wild type oocyte moves specifically to the anterior pole. However, bicoid mRNA particles formed in wild type nurses cells and transferred to kinesin heavy chain mutant oocytes show non-polar localization to the anterior and lateral cortex. Furthermore, we found that the bicoid RNA particles assembled from the oocyte extracts recruit both dynein and kinesin. These observations suggest that kinesin within the oocyte generates anterior-specific bicoid mRNA transport by opposing dynein-dependent movement to the lateral cortex. 927B Differentiation and identity of the Drosophila leg muscles: roles of the FGF pathway and homeodomain transcription factor Ladybird. Tariq Maqbool, Cedric Soler, Teresa Jagla, Jean Philippe Da Ponte, Krzysztof Jagla. INSERM.384, Faculty of Medicine, 28-Place Henri Dunant, 63000-Clermont-Fd, FRANCE. Absract: Leg muscles of the Drosophila display a unique vertebrate-like multi-fibre organization. They are derived from myoblasts associated with the leg imaginal disc, and form a highly stereotyped pattern of ventral and dorsal multi-fibre muscle units attached to the internal tendons. We provide evidence that founder myoblasts represent an obligatory step in the formation of leg muscles, and that the FGF receptor Heartless plays an instructive role in their differentiation This is demonstrated by the expression of read-out of FGF signalling, Dof, which is activated specifically in presumptive founders. Homeobox transcription factor ladybird is expressed in a subset of leg disc myoblasts and FGF is sufficient to ectopically activate ladybird, whose function appears crucial for determining identity of leg versus flight muscles. Overall, we demonstrate that the number of leg founder myoblasts and resulting appendicular fibres is controlled by Heartless transduced FGF signals whereas their identity depends on ladybird. 928C Characterizing the Role of Drosophila LanA in Ommatidial Development. Yu-Chin Lin1,2, Pei-I Tsai1, Cheng-Ting Chien1. 1) Academia Sinica, Institute of Molecular Biology, Taipei, TAIWAN; 2) Institute of Neuroscience, National Yang-Ming University, Taipei, Taiwan. In the development of the Drosophila visual system, establishing correct orientation of each ommatidium is important for their appropriate axonal targeting. To attain their final orientations, ommatidial clusters undergo 90 degree rotation which can be divided into two consequent steps of 45 degree rotation. nemo(nmo), the first identified gene that encodes a MAPK-like protein, promotes the second 45 degree ommatidial rotation. Nemo activity is suppressed by Scabrous (Sca), a signal emitting from morphogenetic furrow to stop the second rotation 27

47th Annual Drosophila Research Conference Late Poster Abstracts at 90 degree position. To identify components that are involved in the rotation process, I take advantage of misrotated phenotypes in the sca and nmo misexpression eyes. By performing EP screening for modifiers of the eye phenotypes, I screened ~1100 independent EP lines, and identified 6 candidate genes that showed genetic interaction with sca and nmo in the mixexpression assays. These candidates include two Rho guanyl-nucleotide exchange factors encoded by trio and RhoGEF, two RNA binding proteins encoded by nanos and FMR1, a zonula adheren component encoded by expanded, and an extracellular matrix (ECM) component Laminin 3/5 encoded by LanA. I further characterized the role of LanA in the ommatidial rotation from adult eye sections and third instar eye imaginal discs. LanA mutants reveal rotation defect which is enhanced by the loss of one copy of nmo. The composition of outer accessory cells in pupal eyes is altered in the LanA mutant and the defect is also enhanced by introducing a copy of nmo mutant. The rotation defects of LanA mutants and its genetic interaction with nmo suggest that the extracellular matrix composed of LamininA may be involved in second rotation and later events. 929A The Control of Ommatidial Death in the Peripehry of the Fly Eye. Andrew Tomlinson, Hui-Ying Lim. Dept Genetics & Development, Columbia Univ, New York, NY. The periphery of the fly eye contains a number of concentrically arranged cellular specializations that are induced by Wingless (Wg) signaling from the surrounding head capsule (HC). The most peripheral of these is the pigment rim (PR) which is a thick layer of pigment cells that lies directly adjacent to the HC, and completely circumscribes the retina. Many of the cells of the PR are derived from presumptive pigment cells that previously surrounded peripheral ommatidia that died by apoptosis. Here we describe the Wg-elicited expression of Snail family transcription factors in the peripheral ommatida that directs their apotosis, and show that expression in a restricted subset of the ommatidial cells is required for the death of the whole unit. These transcription factors control only the death of the peripheral ommatidia; there are no effects upon the other peripheral specializations. We will explain how each specialization appears to be regulated by the Wg-induced expression of specific transcription factors. 930B Functional characterization of CG32062, the Drosophila homologue of human Ataxin-2 Binding Protein. Nagarajan USHA, LS SHASHIDHARA. Centre for Cellular and Molecular Biology, Hyderabad, AP, INDIA. Spinocerebellar ataxia type 2 (SCA2) is a neurodegenerative disease cused by glutamine repeat-expanded Ataxin-2. It is highly conserved in evolution with orthologs in mouse, Caenorhabditis elegans and Drosophila melanogaster. The ataxin-2 activity is modulated by Ataxin-2 Binding Protein 1 (A2BP1), which binds to the Cterminus of Ataxin-2. In an enhancer-trap screen to identify genes that show differential expression between wing and haltere discs, we have identified CG32062, which codes for a RNA-binding protein. It is the closest homologue in Drosophila of human Ataxia-2 binding protein. We have raised antibodies against the bacterially expressed CG32062 protein. In addition, we have generated transgenic flies to knockdown the function of CG32062 using RNAi approach and also transgenic flies to over-express the protein in tissues and developmental stages of our choice. Removal of CG32062 function causes loss of intervein region, mainly between L3-L4 veins of wing blades. The phenotype is often associated with ectopic veins in that region. The Notch pathway specifies the intervein region, while the EGFR pathway specifies veins. The two pathways function antagonistic to each other and thereby draw precise boundaries between vein and intervein regions. Consistently, removal of CG32062 function results in loss of Delta (a ligand required to activate the Notch receptor) and ectopic activation of diphosphorylated form of ERK (MAP kinase), a component of the EGFR pathway. The precise molecular function of CG32062 to positively regulate Notch pathway and/or to negatively regulate EGFR pathway would be discussed.

GAMETOGENESIS AND SEX DETERMINATION 931C

Schpbach1, 2. 1) Howard Houghes Medical Institute; 2) Department of Molecular Biology, Princeton University. Cutoff (cuff) is a female sterile mutation isolated from a large-scale screen. Eggs laid by cuff mutant females display a range of ventralization, consistent with an effect on the Epidermal Growth Factor Receptor (EGFR) pathway. We have found that the cuff mutations affect the accumulation of EGFR ligand, Gurken (Grk), in midstage egg chambers. They also affect germline stem cell maintenance, as well as germline cyst division. We have molecularly cloned cuff. The homologs of cuff have been shown to associate with 5-3 exoribonucleases, and play a role in rRNA processing and transcriptional termination. Currently, we are trying to further characterize the mutation and analyze proteins that interact with Cuff.

Cutoff, a potential exoribonuclease associated protein, is required for Drosophila oogenesis. Yu Chen1, 2, Trudi

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47th Annual Drosophila Research Conference Late Poster Abstracts 932A A role for the Toll-like receptor, 18-Wheeler, in ovarian follicle cell migration. Elizabeth Eldon, Dominic Siler, Cassandra Kleve, Marie Alpuerto. Dept Biological Sci, California State Univ, Long Beach, CA. The ovarian follicle cells have been studied extensively as a model for cell migration, cell signaling and epithelial morphogenesis. 18-wheeler is expressed in a subset of follicle cells that correlate with epithelial migration. To determine the possible role of 18-wheeler expression in these cells, we took advantage of follicle cell-specific FLP expression to generate mitotic clones. Our results show that the posterior migration of epithelial cells that occurs during stage 9, as well as the centripetal migration that occurs during stage 10B, are delayed. The migration of border cells, which do not express 18-wheeler, is unaffected. The eggs deposited by females carrying ovarian clones show altered morphology, consistent both with the expression pattern of 18-wheeler and with the delay in cell migration. Current work is aimed at determining whether a signaling or an adhesion defect underlies the 18wheeler mutant phenotype. 933B Wolbachia preferentially populate the somatic stem cell niche. Horacio M. Frydman1,2, Jennifer Li1,2, Drew Robson1,2, Eric Wieschaus1,2. 1) Howard Hughes Medical Institute; 2) Dept Molecular Biol, HHMI, Princeton Univ, Princeton, NJ. Wolbachia are intracellular bacteria found in the reproductive tissue of all major groups of Arthropods. They are transmitted vertically from the female host to its offspring, in a pattern analogous to mitochondria inheritance. Wolbachia phylogeny, however, does not parallel that of the host, indicating that horizontal infectious transmission has established the bacteria in new hosts during the course of their evolution. The mechanisms for horizontal transfer are largely unknown. We found that Wolbachia cross tissues and infect the germline upon recent introduction into an adult Drosophila female. Based on patterns of enrichment and timing of infection, we conclude that a major route to the germline is through the somatic stem cell niche in the Drosophila germarium. Wolbachia are also highly abundant in the somatic stem cell niche of long term maternally infected hosts, suggesting that continuous infection of the germline from the niche may contribute to its efficient vertical transmission. The investigation of Wolbachia's tropism towards the somatic stem cell niche will deepen our understanding of Wolbachia transmission and of the niche maintaining somatic stem cells. 934C Phosphatidylinositol 4-kinase type II is required for spermatogenesis in Drosophila. Jason Burgess, Ho-Chun Wei, Gordon Polevoy, Julie A Brill. Dept Developmental Biol, Rm 13-401-M, Hospital for Sick Children, Toronto, ON, CANADA, M5G 1L7. The lipid phosphatidylinositol 4-phosphate (PI4P) is synthesized from PI by PI 4-kinases (PI4Ks). PI4P is a key regulator of trafficking through the secretory pathway and is also a precursor of PI 4,5-bisphosphate, which regulates the cytoskeleton, endocytosis/exocytosis and signal transduction. The fwd gene, which is required for male germ cell cytokinesis, encodes a PI 4-kinase (Fwd/PI4KIII). Surprisingly, fwd is not essential, although PI4KIII activity is required for growth and secretion in budding yeast. BLAST analysis revealed that Drosophila contains two additional PI4K encoding genes, PI4KII and PI4KIII. To further investigate the role of PI4P during development, I generated a series of deletions that abolish both somatic and germline specific transcripts of PI4KII. These deletions cause embryonic lethality, suggesting PI4KII is an essential gene. Somatic expression of a PI4KII transgene rescues viability of PI4KII mutant flies. However, the males are sterile and have a defect in maintaining attachment of nuclei to elongated sperm tails at late stages of development. This indicates that both Fwd/PI4KIII and PI4KII are necessary for male fertility. In transgenic flies, a Fwd-GFP fusion protein co-localizes with its lipid product PI4P in spermatocyte Golgi (G.P., H.C.W and J.A.B., unpublished data).To determine the subcellular localization of PI4KII, I generated transgenic flies expressing a PI4KII-RFP fusion protein and found that it colocalizes with PI4P at the Golgi in spermatocytes, at the acroblast in spermatids and in puncta along elongating sperm tails. Thus, PI4KII may participate in vesicle transport during spermatogenesis. 935A lucky luke (luke), a novel gene with a potential function in asymmetric stem cell division. Thomas Fellner1, Allyson Spence1, Cordula Schulz2. 1) Department of Developmental Biology, Stanford University School of Medicine, Stanford, CA; 2) Cold Spring Harbor Laboratories, Cold Spring Harbor, NY. The continuous supply of highly differentiated but short-lived cell types, such as blood, skin, intestinal epithelium, as well as male germ cells, throughout the life of an organism depends on the regulated behavior of adult stem cells. Such cells are relatively undifferentiated and possess a long-term potential to divide and produce two types of daughter cells - those that retain stem cell identity and those that initiate differentiation along a defined lineage. Due to these characteristics, adult stem cells harbor promising potential for regenerative medicine. The Drosophila male germ line is emerging as a powerful model system for the study of the mechanisms that regulate stem cell behavior 29

47th Annual Drosophila Research Conference Late Poster Abstracts in situ, in the context of their normal microenvironment. In a forward genetic screen for viable but male-sterile mutants we identified several new mutants that affect male germ line stem cell behavior. In on of these mutants, lucky luke, we observed that testes lack earlier stages of spermatogenesis while later stages are present, indicating that spermatogenesis is initiated but that stem cell function is not maintained. Further analysis using specific markers revealed that the hub, a cluster of somatic cells at the testis apical tip that functions as a component of the germ line stem cell niche, is retained in the mutant and surrounded by stem cells. This indicates that male germ line stem cells in this mutant are initially functional but loose their capability to produce differentiating daughters over time. The phenotypic and molecular characterization of this new gene will be the topic of my presentation.

ORGANOGENESIS 936B Regulation of Drosophila Friend of GATA gene, u-shaped, during hematopoiesis: A direct role for Serpent and Lozenge. Selen Muratoglu1, Betsy Garratt1, Kristy Hyman2, Kathleen Gajewski2, Robert A. Schulz2, Nancy Fossett1. 1) Center for Vascular and Inflammatory Diseases and the Department of Pathology, University of Maryland School of Medicine, Baltimore, MD 21201; 2) Department of Biochemistry and Molecular Biology, Graduate Program in Genes & Development, The University of Texas M. D. Anderson Cancer Center, Houston, TX 77030. Friend of GATA proteins interact with GATA factors to regulate development in a variety of tissues. To better understand the transcriptional control of this important gene family, we analyzed cis- and trans-regulation of the Drosophila Friend of GATA gene, u-shaped. Using overlapping genomic fragments driving tissue-specific reportergene (lacZ) expression, we showed that most of the embryonic cis-regulatory elements are located within the 7.4 kb region upstream of the transcription start site. We also identified two minimal hematopoietic enhancers in this region. One was active in all classes of hemocytes; whereas, the other was active in hemocyte precursors and plasmatocytes only. The GATA factor, Serpent, directly regulated the activity of both enhancers. However, activity in the crystal cell lineage not only required Serpent but also the RUNX homologue, Lozenge. These findings represent the first demonstration of GATA and RUNX regulation of Friend of GATA gene expression. In addition, we analyzed cis-regulation of u-shaped expression in the lymph gland and identified similarities and differences between regulatory strategies used during embryonic and lymph gland hematopoiesis. The results of these studies provide information to further analyze the regulation of this important conserved gene family. 937C Expression and mutual repression of tinman and the Dorsocross genes establishes diversified cardiac cell identities in the dorsal vessel. Ingolf Reim1, Stphane Zaffran2, Manfred Frasch1. 1) Mol., Cell & Dev. Biology, Mount Sinai School of Medicine, New York, NY; 2) Dev. Biology Inst. of Marseille-Luminy, Marseille, France. The conserved NK-homeobox gene tinman (tin) is essential for the specification of the cardiac and visceral muscle progenitors in the early dorsal mesoderm. These functions depend on the twist- and dpp-dependent early phases of tin expression. Later, the expression of tin is maintained in cardiac cells during cardiac maturation and differentiation; however, the function of tin in these cells has not been defined. To investigate the role of tin during cardiac patterning and differentiation, we generated tin mutants in which tin expression is rescued within the early and dorsal mesoderm, but not within cardiac cells. Analysis of these animals shows that myocardial cells and dorsal vessels can form in the absence of cardiac Tin activity, and the animals can survive into adult stages. However, embryos in which Tin is missing from cardiac cells show severe disruptions in myocardial cell diversification. Importantly, our study shows that the normal expression of Tin in four of the six bilateral cardioblasts within each hemisegment of the heart allows these cells to adopt a generic myocardial cell fate as opposed to a fate as inflow tract (ostial) cells. One feature of ostial cells is the expression of the T-box transcription factors Dorsocross (Doc1-3). We show that Tin acts as a repressor of Doc expression in the presumptive generic myocardial cells. Notably, the repressive activity of Tin is conserved in its mouse ortholog Nkx2.5, as shown by misexpression studies. Conversely, we demonstrate that maintained Doc activity acts to repress tin expression in ostial cells, which reinforces an initial tin-repressing activity provided by the COUP-TF-related orphan nuclear receptor transcription factor Seven-up (Svp). Ultimately, the mutually exclusive expression of Tin and Doc in these two types of myocardial cells leads to the differential expression of certain differentiation genes and their characteristic morphological and functional distinction.

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47th Annual Drosophila Research Conference Late Poster Abstracts 938A Positive and Negative Control of Muscle Differentiation. Michael Taylor, David Liotta. Sch Biosciences, Cardiff Univ, Cardiff, United Kingdom. A balance of promoting and restraining influences is thought to control when and where cells differentiate during development. We are analysing this issue during muscle differentiation. To date there is very little in vivo information in any species on molecules that restrain the differentiation of muscle. However, in a screen we identified a novel gene, Dmeso17A, which our results indicate is part of a restraining mechanism. Dmeso17A expression rapidly declines in the somatic mesoderm as embryonic muscle differentiates. However, it persists in the adult muscle precursors that are set aside in the somatic mesoderm and which remain undifferentiated at this stage. Consistent with this expression pattern we found that Dmeso17A over-expression inhibits muscle differentiation. This effect is suppressed by over-expression of the Dmef2 transcription factor, the key promoter of muscle differentiation. These results suggest that Dmeso17A is an inhibitor in the Dmef2-regulated pathway of muscle differentiation. We explored the Dmeso17A loss-of-function phenotype through dominant negative and RNAi constructs. These result in aberrant somatic muscle differentiation. How might Dmeso17A be working? It is found in the nucleus and we are analysing the relationship between it and possible interacting genes during muscle differentiation. Our model is that Dmeso17A down-regulates Dmef2, and that it down-regulates Dmef2 activity rather than Dmef2 expression. In support of this we found that Dmeso17A inhibits the expression of a range of Dmef2 target genes, but has no effect on the expression of Dmef2 itself. Taken together, our results suggest that Dmeso17A is a novel component of a muscle differentiation switch, and is part of a mechanism that holds cells in an undifferentiated state.

NEUROGENETICS AND NEURAL DEVELOPMENT 939B The F-actin/Microtubule crosslinker Shot is a platform for Krasavietz-mediated translational regulation of midline axon repulsion. Seungbok Lee1, Seongsoo Lee1, Peter Kolodziej2. 1) School of Dentistry, Seoul National Univ, Seoul, KOREA; 2) Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN, USA. Axon extension and guidance require a coordinated assembly of F-actin and microtubules and regulated translation in the growth cone. The molecular basis of how the translation of mRNAs encoding guidance proteins could be closely tied to the pace of cytoskeletal assembly is poorly understood. Previous studies showed that Short Stop (Shot) mediates direct interactions between F-actin and microtubules required for motor and sensory axon extension in the Drosophila embryo. Here, we provide biochemical and genetic evidence that Shot functions in neurons together with a novel translation inhibitor, Krasavietz (Kra), to steer longitudinally directed CNS axons away from the midline. Kra can bind directly to the C-terminal domain of Shot. Mutations in shot and kra lead to weak robo-like phenotypes and dominantly enhance each others midline phenotypes. shot and kra genetically interact with slit. We demonstrate that Kra interacts with the translation machinery and inhibits translation in vitro. These data suggest that Shot-Kra interaction controls the translation of mRNAs encoding proteins necessary for the Slit/Robo response of the growth cone. We propose previously unknown role of Shot as a direct physical link between translational regulation and cytoskeleton reorganization. 940C Highly ordered assembly of retinal axons and their synaptic partners is regulated by Hedgehog/Single-minded in Drosophila visual system. Daiki Umetsu, Satoshi Murakami, Makoto Sato, Tetsuya Tabata. Dept Cell Biol, IMCB Univ Tokyo, Tokyo, JAPAN. In development of the Drosophila visual center, photoreceptor cells extend their axons (R axons) to the lamina ganglion layer and trigger proliferation and differentiation of synaptic partners (lamina neurons) by delivering the inductive signal, Hedgehog (Hh). This inductive mechanism helps to establish an orderly arrangement of connections between the R axons and lamina neurons, termed a retinotopic map because it results in positioning the lamina neurons in close vicinity to the corresponding R axons. We found that the bHLH-PAS transcription factor Single-minded (Sim) is induced by Hh in the lamina neurons and is required for the association of lamina neurons with R axons. In sim mutant brains, lamina neurons undergo the first step of differentiation but fail to associate with R axons. As a result, lamina neurons are set aside from R axons. The data reveal a novel mechanism for regulation of the interaction between axons and neuronal cell bodies that establishes precise neuronal networks.

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47th Annual Drosophila Research Conference Late Poster Abstracts 941A Screening of novel genes related to neuronal apoptosis mediated by dAPP-BP1 pathway in Drosophila melanogaster. Songhee Kim, Kiyoung Kim, Joohee Lee, Jongwoo Ryu, Jeongbin Yim. School of Biological Sciences, Seoul National University, seoul, Seoul, KOREA. Alzheimers disease (AD) is a progressive dementia. Amyloid precursor protein (APP) is the source of aggregated amyloid and linked to the early onset of AD. APP-BP1 was first identified as an APP carboxy-terminal binding protein. APP-BP1 also fulfill E1-like function to activate the small-ubiquitin-like protein NEDD8. It is known that expression of APP-BP1 in dividing cells drives the cell cycle through the S-M checkpoint and overexpression of APP-BP1 causes apoptosis in primary neurons. In Drosophila, loss of dAPP-BP1 (Drosophila homolog of APPBP1) cause neuronal apoptosis like human APP-BP1. However the exact mechanism from APP-BP1 to apoptosis is not known yet. To investigate mechanism of neuronal apoptosis regulated by APP-BP1 and find novel genes related in this process, we performed EMS (ethane methyl sulfonate) mutagenesis in Drosophila. Through EMS mutagenesis, we screened drosophila that link dAPP-BP1 pathway and apoptotic process. So far we have found 13 candidates that rescue the lethality induced by dAPP-BP1 null mutation. The novel function of these genes that take part in the apoptosis pathway induced by loss of APP-BP1 is currently under investigation. 942B Confocal analysis of the Drosophila mutant small mushroom bodies (smu) reveals an abnormal cell number and axonal pattern. Brian Dunkelberger, Christine Serway, Steven de Belle. University of Nevada, Las Vegas, Las Vegas, NV.89154-4004. Mushroom bodies are paired neuronal assemblies in the insect brain that have been implicated in olfactory memory integration. Histological preparation of the mutant small mushroom bodies (smu) reveals a severely reduced mushroom body calyx volume. In these experiments, the reason for this reduction is shown to be caused by a reduced cell number which is reflected in abnormal axonal architecture. 943C Analysis of the role of Drosophila Phosphotyrosyl phosphatase activator (D-Ptpa) in peripheral nervous system development. Michelle Brooks-Pigott1,2,3, Eddy Van Daele4, Veerle Janssens5, Christine Van Hoof5, Jozef Goris5, Koen Norga1,2,6, Patrick Callaerts1,2,3. 1) Laboratory of Developmental Genetics, Flanders Institute of Biotechnology (VIB); 2) Center for Human Genetics, University of Leuven, Leuven, BE; 3) University of Houston, Department of Biology and Biochemistry, Houston, Texas (former affiliation); 4) University of Sussex, School of Biological Sciences, Brighton, UK; 5) University of Leuven, Afdeling Biochemie, Faculteit Geneeskunde, Departement Moleculaire Celbiologie, Leuven, BE; 6) Childrens Hospital, University of Leuven, Leuven, BE. Reversible phosphorylation is a key event in many biological processes and is controlled by the activities of kinases and phosphatases. Protein phosphatase 2A (PP2A) comprises a family of serine/threonine phosphatases that has been shown to function as a major cellular phosphatase involved in many fundamental biological processes. Furthermore, dysfunction of PP2A has been linked to neurodegeneration, neurodevelopmental disorders and cancer. However, very little is known about the role of PP2A during nervous system development. During peripheral neurogenesis, sensory organ precursor cells (SOPs) are defined within proneural fields. Both the Notch and EGFR signaling pathways have been identified as key regulators of SOP selection. Drosophila phosphatase 2A phosphatase activator (D-Ptpa) is a cofactor of PP2A, specifically stimulating the phosphatase activity of the protein in a reversible manner. Here, we show that loss of D-Ptpa results in bristle loss, transformation of bristle type, ectopic neurons and a loss of SOPs. This suggests that D-Ptpa plays a role at multiple stages in SOP development. In addition, we observe interaction of D-Ptpa with twins (tws) as well as with microtubule star (mts), members of the B and C subunit families of PP2A, respectively. Finally, we find that loss of D-Ptpa interacts dominantly with Notch and EGFR pathway mutants. We hypothesize that D-Ptpa, through its interaction with PP2A, is involved with these pathways in SOP development.

NEURAL PHYSIOLOGY AND BEHAVIOR 944A The Drosophila Mutant oblivious Is Defective In Nociception. Lixian Zhong4, W. Daniel Tracey1,2,3. 1) Dept. of Anesthesiology; 2) Dept. of Cell Biology; 3) Dept. of Neurobiology; 4) Dept. of Pharmacology, Duke University, Durham, NC. In a genetic screen for abnormal larval behavioral responses to noxious heat, we have identified a mutant which we named oblivious. In oblivious mutant larvae the behavioral response to noxious heat is severely impaired. The mutant line contains a single EP element insertion that is located between two candidate genes that may be 32

47th Annual Drosophila Research Conference Late Poster Abstracts responsible for the mutant phenotype. One of the genes encodes a G protein coupled receptor (GPCR) whose transcriptional start site is located approximately two kilobases from the EP element insertion site. This receptor, CG10823, is the Drosophila orthologue of the human neuropeptide FF (NPFF) receptor that may play a role in modulation of pain in vertebrates. The other candidate gene is the non-coding RNA hsr-omega that is located only about 150 bp from the insertion site. By quantitative RT-PCR, transcription of CG10823 is upregulated in the oblivious mutant strain. GAL4 enhancer traps in the first intron of CG10823 shows expression in a small number of neurons in the larval brain but not in multidendritic neuron nociceptors. In contrast, an enhancer trap within the hsromega transcription unit does show expression in multidendritic neurons, as well as expression in complex central nervous system circuitry. We will present the results of genetic experiments that are in progress to determine whether the EP-element insertion causes the mutant phenotype or if the phenotype is due to an unlinked mutation elsewhere on the chromosome. 945B A Sleep-promoting Role for the Drosophila Serotonin Receptor 1A. Quan Yuan1, William Joiner2, Amita Sehgal1, 2. 1) Howard Hughes Medical Institute; 2) Center for Sleep and Respiratory Neurobiology, University of Pennsylvania, School of Medicine, Philadelphia, PA. Although sleep is an important process essential for life, its regulation is poorly understood. The recently developed Drosophila model for sleep provides a powerful system to genetically and pharmacologically identify molecules that regulate sleep. Serotonin is an important neurotransmitter known to affect many behaviors, but its role in sleep remains controversial. We generated or obtained flies with genetically altered expression of each of three Drosophila serotonin receptor subtypes, d5-HT1A, d5-HT1B and d5-HT2, and assayed them for baseline sleep phenotypes. The data indicated a sleep-regulating role for the d5-HT1A receptor. d5-HT1A mutant flies had short and fragmented sleep, which was rescued by expressing the receptor in adult mushroom bodies, the structure associated with learning and memory in Drosophila. Neither the d5-HT2 receptor, nor the d5-HT1B receptor, which was previously implicated in circadian regulation, had effects on baseline sleep, indicating that serotonin affects sleep and circadian rhythms through distinct receptors. Elevating serotonin levels, either pharmacologically or genetically, enhanced sleep in wild type flies. In addition, serotonin promoted sleep in two short-sleep mutants suggesting that it can compensate for some sleep deficits. These data suggest that serotonin promotes sleep in Drosophila. They also link the regulation of sleep behavior by serotonin to a specific receptor in a distinct region of the fly brain. 946C Classical reward conditioning in Drosophila melanogaster. Kyung-An Han, Young-Cho Kim, Hyun-Gwan Lee. Department of Biology, Pennsylvania State Univ, University Park, PA. Negatively reinforced olfactory conditioning has been widely employed to identify learning and memory genes, signal transduction pathways and neural circuitry in Drosophila. In an effort to delineate the molecular and cellular processes underlying reward-mediated learning and memory, we developed a new assay for positively reinforced olfactory conditioning. Flies were involuntarily exposed to appetitive unconditioned stimulus sucrose along with conditioned stimulus odor during training and their preference to the odor previously associated with sucrose was measured to assess learning and memory capacities. After one training session, wild-type Canton S flies displayed reliable performance, which was enhanced after two training cycles with 1 or 15 min inter-training interval. Also, higher performance scores were obtained with increasing sucrose concentration. Memory in Canton S flies was slowly decayed when measured at 30 min, 1 hr and 3 hr after training; whereas, it was significantly declined at 6 hr, which was maintained at 12 hr post-training. When the learning mutant, t?h, deficient in octopamine was challenged, they exhibited poor performance, validating the utility of this assay. Moreover, oamb (octopamine receptor) mutants revealed impaired learning in this paradigm, which was rescued by transgenic expression of OAMB in the mushroom bodies. Considering tremendous resources of genetic and transgenic mutants and tools available in Drosophila, the new reward conditioning described here will provide a useful tool to elucidate the neurobiological basis of learning and memory. Additionally, this reward olfactory conditioning represents classical conditioning as does negatively reinforced olfactory conditioning. Thus, comparative analysis of learning and memory mutants in two assays will help dissect molecular and cellular processes common or discrete to US information used in conditioning. 947A Drosophila Dorsal Paired Medial neurons provide a general mechanism for prolonging memory. Michael J Krashes, Alex C Keene, Jessica A Bernard, Scott Waddell. Neurobiology, UMass Medical School, Worcester, MA. Memories are formed, stabilized in a time-dependent manner, and stored in neural networks. In Drosophila, retrieval of aversive and appetitive odor memories depend on output from mushroom body (MB) neurons consistent 33

47th Annual Drosophila Research Conference Late Poster Abstracts with the idea that both types of memory are represented there. Dorsal Paired Medial (DPM) neurons innervate the mushroom bodies and DPM neuron output is required for the stability of aversive odor memory. Here we show that stable appetitive odor memory is also DPM neuron dependent. DPM neuron expression of amnesiac (amn) in amn mutant flies restores wild-type memory. In addition, disrupting DPM neurotransmission between training and testing abolishes appetitive odor memory, like it does with aversive memory. We further examined DPM-MB connectivity by overexpressing a DScam variant that selectively disrupts DPM neuron projections to the MB alpha, beta and gamma lobes. Prime lobe projecting DPM neurons are capable of stabilizing aversive and appetitive odor memory implying very similar circuit requirements for both forms of memory. Therefore our results suggest that the fly employs the local DPM-MB circuit to stabilize aversive and appetitive odor memories and that stable aspects of both forms of memory likely reside in mushroom body prime lobes. 948B Multiple memories of Drosophila sugar reward learning. Andreas S. Thum, Martin Heisenberg, Hiromu Tanimoto. Department of Genetics and Neurobiology, University of Wrzburg, Biozentrum am Hubland, Germany. Drosophila melanogaster forms associative olfactory memories with electric shock as an aversive reinforcer or sucrose reward as appetitive reinforcer. The Kenyon cells of the mushroom bodies have been shown to contain engrams of both memories. Several lines of evidence from honeybee, moth and fly have suggested another site, the antennal lobe to house neuronal plasticity underlying appetitive olfactory memory. Here we describe a new engram of olfactory memory in the Drosophila projection neurons by restoring Rutabaga adenlylate cyclase (rutAC) activity specifically in these cells. Expression of wild-type rutabaga in the projection neurons fully rescued the defect in rewarded memory, but not in aversive electric shock memory. We found no difference in the stability of the appetitive memories rescued either in projection neurons or Kenyon cells. Our results suggest that olfactory memory may be conserved between insect species even at the circuit level. 949C Protein Phosphatase 1 Regulates the Stability of Drosophila Circadian Clock Proteins. Yanshan Fang, Sriram Sathyanarayanan, Amita Sehgal. Howard Hughes Medical Institute, Department of Neuroscience, University of Pennsylvania School of Medicine, Philadelphia, PA 19104. Circadian rhythms in Drosophila is maintained by a feedback loop, in which the protein products of two major clock genes period (per) and timeless (tim) inhibit their own transcription by repressing transcriptional activators dCLOCK (dCLK) and CYCLE (CYC). Timely degradation of PER and TIM relieves the inhibition at late night, and therefore sustains rhythmic RNA and protein levels of per and tim. However, PER and TIM still oscillate with a circadian rhythm in the absence of rhythmic transcription, indicating posttranslational regulations of clock proteins. Several clock kinases including DOUBLETIME, SHAGGY, and casein kinase 2 have been identified in Drosophila, and phosphorylation by these kinases regulates PER and TIM stability and subcellular translocation. Recently, we showed protein phosphatase 2A (PP2A) promotes the stability and the nuclear accumulation of PER through dephosphorylation. We now report another major protein serine/threonine phosphatase, protein phosphatase 1 (PP1), is involved in regulating the stability of PER and TIM. In Drosophila S2 cells, inhibition of PP1 by overexpression of the nuclear inhibitor of PP1 (NIPP1) or incubation with PP1 dsRNA reduces the protein levels of transfected TIM and PER. Co-immunoprecipitation assays showed that PP1 binds to TIM and PER in transfected S2 cells. Moreover, flies overexpressing NIPP1 in clock cells have lower TIM protein levels and exhibit longer circadian period. Currently, experiments are under way to determine whether phosphorylation states and nuclear accumulation of TIM and PER are affected by PP1 and to identify the relevant PP1 isoform for the circadian effects. Together, these studies will further our understanding of the posttranslational mechanisms that underlie circadian clock function in Drosophila. 950A FOXO and insulin signaling regulate the sensitivity of circadian clock to oxidative stress. Xiangzhong Zheng, Amita Sehgal. Dept. Neurosci, 232 Stemmler Hall, Univ. Pennsylvania Med. Sch./HHMI, Philadelphia, PA 19104. Oxidative stress affects numerous cellular processes. We show here that circadian rhythms in Drosophila are sensitive to oxidative stress, particularly in the absence of the FOXO protein. When exposed to oxidative stress, wild type flies showed attenuated clock gene cycling in peripheral tissues, while foxo mutants also lost behavioral rhythms driven by the central clock. FOXO was expressed predominantly in the fat body. Insulin signaling was elevated in foxo mutants as indicated by increased levels of the putative insulin binding protein, acid labile subunit (als), increased nuclear localization of the insulin receptor,and loss of behavioral hyperactivity in response to restricted feeding. In addition, over-expression of components of the insulin signaling pathway in the fat body also increased susceptibility of the central clock to oxidative stress. Behavioral rhythms declined rapidly with age in foxo mutants. Together these data demonstrate a non-cell-autonomous mechanism that affects central clock function, and provide a link between insulin signaling, oxidative stress, circadian rhythms and aging. 34

47th Annual Drosophila Research Conference Late Poster Abstracts

EVOLUTION AND QUANTITATIVE GENETICS 951B Evolution Through the Eyes of a Fly. Margarita Ramos, Alistair McGregor, David Stern, Peter Grant. Ecology and Evolutionary Biology, Princeton University, Princeton, NJ. Phenotypic variation is the essence of the process of evolution. In order to generate a comprehensive understanding of how evolution proceeds we need to characterize both proximate and ultimate causes generating phenotypic variation. A combination of both laboratory and field studies of closely related species might allow us to identify genetic changes responsible for phenotypic differences and the selective forces that brought them forth. The eye is a complex morphological structure that has diversified into a varied array of types to suit the lifestyle of its bearer making it a suitable trait to understand evolutionary processes. Within the melanogaster species subgroup, Drosophila mauritiana has strikingly larger and differently shaped eyes (about 30% more ommatidia) than its sibling species Drosophila simulans. In this ongoing project, we seek an integrative approach to understand the evolution of this character by examining both its proximate (i.e. genetic basis, development), and its ultimate causes (selective forces in nature). Here we present data on the identification of genomic regions responsible for differences in eye size between D. simulans and D. mauritiana. A previous study identified a region close to the forked locus as significantly associated with this trait. We confirmed this conclusion through a rough QTL mapping analysis, and have further refined the mapping of a locus responsible for approximately 50% of the variation in this trait using primarily meiotic recombination mapping.

IMMUNE SYSTEM AND CELL DEATH 952C Collier/Knot is required for PSC formation and control of blood cell homeostasis. Alain Vincent1, Joanna Krezmien1, Rami Makki1, Laurence Dubois1, Jean Michel Ubeda2, Marie Meister2, Michle Crozatier1. 1) Centre de Biologie du Dveloppement, CNRS/Universit, Toulouse, France; 2) Institut de Biologie Molculaire et Cellulaire, CNRS, Strasbourg, France. We have previously shown that Collier/Knot (Col), the Drosophila ortholog of mammalian Early B-Cell Factor (EBF) controls the production of lamellocytes, the specific cellular immune response to wasp parasitization. Col is expressed very early during ontogeny of the lymph gland (the larval hematopoietic organ) and this expression becomes quickly restricted to a few cells prefiguring the so-called Posterior Signalling Center (PSC) first described by U. Banerjees lab. We initially showed that Col endows PSC cells with the capacity to relay an instructive signal and orient hematopoietic precursors towards the lamellocyte fate in reponse to parasitization (Crozatier et al, PLOS Biol, 2004. We have now extended these studies to that Col is required for PSC formation and that the PSC plays a general, critical role in maintaining blood cell homeostasis in absence of, and response to a immune challenge. The signalling pathways involved in the communication between PSC cells and the multipotent hematopoietic progenitors in these different conditions will be discussed. 953A Establishing the molecular events during pattern recognition of Gram-positive bacteria in Drosophila innate immunity. Lihui Wang1, Alexander Weber2, Sergio Filipe3, Nicholas Gay2, Petros Ligoxygakis1. 1) Genetics Unit, Department of Biochemistry, University of Oxford, South Parks Rd OX1 3QU, Oxford UK; 2) Department of Biochemistry University of Cambridge, 80 Tennis Court Road, CB2 1GA Cambridge UK; 3) Instituto de Tecnologia Qumica e Biolgica / Universidade Nova de Lisboa, Av. da Repblica (EAN), Apartado 127, 2781-901 Oeiras, Portugal. Genetic evidence indicates that Drosophila defence against Gram-positive bacteria is mediated by two putative pattern recognition receptors acting upstream of Toll. Mutations in either Gram-Negative Binding Protein 1 (GNBP1) or Peptidoglycan Recognition Protein-SA (PGRP-SA) compromise Toll-dependent response to infection. In the present we report that recombinant GNBP1 is able to bind and hydrolyse Gram-positive peptidoglycan (PG). PGRP-SA is able to interact with GNBP1 and this interaction is significantly enhanced in the presence of GNBP1hydrolysed PG or highly purified PG fragments. PGRP-SA binds to PG fragments and this interaction depends on the polymerisation status of the muropeptides, pointing to constraints in the number of PGRP-SA molecules bound for signalling initiation. We propose a model whereby GNBP1 presents a more palatable form of PG for sensing by PGRP-SA and that physical interaction between these proteins in the presence of PG results in downstream signalling. 35

47th Annual Drosophila Research Conference Late Poster Abstracts 954B Dmp53 activates the Hippo pathway to promote cell death in response to DNA damage. Julien Colombani, Cedric Polesello, Filipe Josue, Nicolas Tapon. Apoptosis and Proliferation Control Lab, Cancer Research UK, London Institute, United Kingdom. The Hippo (Hpo) pathway comprises the kinases Hpo and Warts (Wts or LATS), the adaptors Salvador (Sav) and Mob1 as a tumour suppressor (Mats), the cytoskeletal proteins Expanded and Merlin, and the transcriptional cofactor Yorkie (Yki). This pathway has been shown to restrict cell division and promote apoptosis during development in Drosophila. Though we understand some of the events downstream of the Hpo pathway, its mode of activation remains mysterious. In the present study, we show that Hpo can be activated by Ionizing Radiations (IR) in a Dmp53 (Drosophila melanogaster p53)-dependent manner, and that Hpo is required for the cell death response elicited by IR or Dmp53 ectopic expression. This suggests that Hpo is necessary for the apoptotic response to DNA damage and provides the first input for Hpo activation. Since mammalian components of the Hpo pathway have been suggested to behave as tumour suppressors, we propose that mutations in its members may protect tumour cells against the apoptotic effects of IR. 955C Alteration of gravitational force affects the immune response. Katherine A Taylor1, Alexander T Kloehn1, Patrick M Fuller2, Kathleen M Beckingham3, Charles A Fuller2, Deborah A Kimbrell1. 1) Molecular & Cellular Biology, University of California, Davis, CA; 2) Neurobiology, Physiology & Behavior, University of California, Davis, CA; 3) Dept. Biochemistry and Cell Biology, Rice University, Houston, TX. Immune dysfunction has serious consequences and is a major concern for functioning in the space environment. An understanding of the relationship between gravitational forces and immune function is required. In order to advance this understanding, we have initiated Drosophila as a model of immune function under conditions of altered gravity. In the current studies, we are examining parameters of immune function under conditions of hypergravity after infection with fungus, Beauveria bassiana, or bacteria, E. coli. A critical feature is the ability to compare wild type flies to both immune response mutants and to gravity sensing mutants. Our data show that exposure to hypergravity changes the survival of infected flies. Specifically, flies show a significant increase (P <0.05) in survival rates at 4g versus 1g. An increase in survival rate is consistent with either the infecting organism being weaker and/or the flies being stronger in response. By comparing these results to gravity sensing mutants, the relative roles of gravity, host and pathogen can begin to be discerned. We thus tested yuri mutants, named after the Russian astronaut Yuri Gagarin and isolated as having a defective gravitaxis response, for their survival after infection and exposure to 4g hypergravity. The UAS-yuri rescue stock showed the same increase in survival as the other stocks tested, but yuri mutants showed no change in survival at 4g compared to 1g. Thus, the data indicate that gravity has an effect on the host response. Subsequent tests are in progress to further determine the relative effects of gravity on the functions of the host and infecting organism.

TECHNIQUES AND GENOMICS 956A An efficient genetic screen in Drosophila to identify nuclear-encoded genes with mitochondrial function. Albert Cespedes1,2, T. S. Vivian Liao1,2, Gerald B. Call1,2, Preeta Guptan1, Jamie Marshall1, Edward Owusu-Ansah1, Sudip Mandal1, Q. Angela Fang1, Kevin Yackle1, Gelsey L. Goodstein1, William Kim1, Utpal Banerjee1. 1) Mol., Cell and Dev. Biology, UCLA, Los Angeles, CA; 2) These authors contributed equally to this work. We have recently published our findings on tenured which is a mutation in cytochrome c oxidase subunit Va of Complex IV of the mitochondrial electron transport chain. This mutation allows cells in the developing eye to undergo a number of cell divisions, but eventually causes a checkpoint in the G1-S phase of the cell cycle. This cell cycle checkpoint is caused by a reduction in ATP levels to about 40% of wild type. Since we identified this mutation by an adult glossy eye, and a lack of BrdU incorporation in the second mitotic wave in the third instar larval eye disc, we undertook a screen to identify other genes on chromosome arm 3R that give a similar phenotype. Out of 75,000 mutant chromosomes analyzed, there were 90 mutants identified, comprised of 46 complementation groups, that gave an adult glossy eye. These 46 groups were all subsequently assayed for BrdU incorporation and stained for photoreceptor development using the neuron-specific ELAV antibody. Based on these two criteria, the

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47th Annual Drosophila Research Conference Late Poster Abstracts mutants were categorized into 4 classes: both wild-type BrdU and ELAV (10), wild-type BrdU and defective ELAV (3), abnormal BrdU and defective ELAV (10), and abnormal BrdU and wild-type ELAV (23). The latter category of mutants were followed up as part of this screen to identify other genes involved in this G1-S checkpoint. Five of these genes have been molecularly characterized to date and all represent proteins involved in mitochondrial function. CG6022, a homolog of cytochrome c heme lyase (cchl); CG9613, a homolog of coenzyme Q biosynthesis protein 2 (coq2); CG10092, a homolog of arginyl tRNA ligase (atl); ATP synthase delta subunit (ATPsyn-); and glutamyl-tRNA amidotransferase A (gatA). Combining this with our previous findings demonstrates the effectiveness of our screen to identify genes with mitochondrial function. 957B Insect Sequencing at the Baylor College of Medicine Human Genome Sequencing Center. Stephen Richards1, Yue Liu1, Kim C. Worley1, Erica Sodergren1, Steven E. Scherer1, Catherine M. Rives1, Susan Brown2, Richard Beeman3, John H. Warren4, Donna M. Muzny1, George Weinstock1, Richard A. Gibbs1. 1) Human Genome Sequencing Center, Baylor College of Medicine, Houston TX; 2) Division of Biology, Kansas State University, Manhattan, KA; 3) USDA, Biological Research Unit, 1515 College Ave, Manhattan KA; 4) Department of Biology, University of Rochester, NY. Genome sequences greatly facilitate biological research - especially in model organisms with facile biology. Having sequenced a portion of D. melanogaster, all of D. pseudoobscura and the honeybee, the Human Genome Sequencing Center (HGSC) is currently annotating Tribolium castaneum (red flour beetle), sequencing the parasitic wasp Nasonia vitripennis and sibling species N. giraulti and N. longicornis, and the pea aphid (Acyrthosiphon pisum). The Tribolium genome comprises of 152Mb of assembled sequence with 70% anchored to the 10 chromosomes. Automated gene prediction methods produce ~ 14,000 gene models, of which ~ 9,500 gene models are supported by transcribed sequence, or protein homology. Effective RNAi inhibition allows immediate progression from sequence to experiment. We present an example showing differences in pair-rule genes between D. melanogaster and T. castaneum. Tribolium hairy does not function in segmentation. However, odd-skipped, a secondary pair-rule gene in Drosophila, functions as a primary pair-rule gene in Tribolium. Nasonia is in progress at the HGSC. The sequence of the three Nasonia species creates a new paradigm for cloning quantitative trait loci. These species, separated by Wolbachia infection causing sperm egg incompatibility, can interbreed after antibiotic treatment. Male haploidy in the hymenoptera allows polygenic phenotypic differences between the species (natural or selection induced) to be rapidly mapped to genes. 6X sequence coverage of N. vitripennis genome is in the NCBI trace archive, with assembly imminent. 1X sequencing of sibling species is currently underway, to produce the required polymorphism maps. Finally, we present progress of pea aphid sequencing at the HGSC. 958C Use Drosophila as A Model System to Identify the Function of Human Genes. Yung-Heng Chang1,2, Y. Henry Sun2. 1) Graduate Institute of Life Sciences, National Defense Medical Center, Taipei 114, Taiwan, Republic of China; 2) Institute of Molecular Biology, Academia Sinica, Taipei 115, Taiwan, Republic of China. After the finish of human genome sequencing, how to annotate and identify the function of one human gene becomes an interesting and challenging field. Targeted expression of many human genes in Drosophila were found to causes some morphological phenotypes, suggesting that the human genes are interfering some conserved molecular mechanisms. Therefore, Drosophila provides an excellent in vivo system to study the function of these novel human genes. In collaboration with seven fly labs in Taiwan, we selected one hundred human genes whose expression level changed significantly in hepatoma cells and also changed in cell cycle. Using the GAL4-UAS system, we expressed these transgenes in various fly tissues in wild type or sensitized genetic backgrounds and screened for specific phenotypes. In our study, these human genes were induced to express in the eye by several eye specific GAL4 and in the whole organism by tub-GAL4. We also test the genetic interaction of these human genes with eyg and upd in which regulate cell proliferation during eye development. Several genes were identified to give interesting phenotypes. These will be presented.

DROSOPHILA MODELS OF HUMAN DISEASES 959A An inducible Drosophila cell model to study polyglutamine disease. Wing Man Chan1,2, Pang Chui Shaw2, Edwin HY Chan1,2. 1) Laboratory of Drosophila Research, The Chinese University of Hong Kong, HKSAR, China; 2) Molecular Biotechnology Programme, Department of Biochemistry, The Chinese University of Hong Kong, HKSAR, China. Polyglutaminopathies define a group of neurodegenerative diseases characterized by the presence of an elongated polyglutamine (polyQ) domain in the disease proteins, owing to the expansion of existing CAG 37

47th Annual Drosophila Research Conference Late Poster Abstracts trinucleotide repeats in the corresponding disease genes. Common cellular pathologic features of polyglutaminopathies include progressive neuronal degeneration and formation of insoluble intracellular protein aggregates. The observed pathological similarities among different polyQ disorders support the existence of shared molecular disease pathways. We established an inducible Drosophila cell model to study polyQ disease. The BG2-c6 neuronal cell line was stably transfected with various lengths of GFP-polyQ constructs. Western blotting, filter trap assay and fluorescence microscopy have been performed to characterize the effects of polyQ protein expression on BG2-c6 cells. In particular, cytoplasmic and nuclear distributions of polyQ protein aggregates were studied in detail. This is because formation of polyQ aggregates is correlated with polyQ-induced toxicity. We found that sodium dodecyl sulphate (SDS)-insoluble polyQ aggregates accumulated in the nucleus in a Q-length and timedependent manner, whereas cytoplasmic polyQ aggregates remained SDS-soluble. The difference of aggregate solubility properties in different subcellular compartments may explain the well-documented transcriptional dysfunction in polyQ-induced toxicity. 960B A NEW DROSOPHILA MODEL FOR SCA7 POLYGLUTAMINE DISEASE DEMONSTRATES REVERSIBLE NEUROTOXICITY AND ALLOWS QUANTITATIVE SCREENING FOR PHENOTYPIC MODIFIERS. Lasbleiz Christelle1, Monnier Vronique1, Latouche Morwena2, Brice Alexis2, Stevanin Govanni2, Tricoire Herv1. 1) Institut Jacques Monod, Paris, FRANCE; 2) Institut des Neurosciences, Paris, France. We have created a conditional model of spinocerebellar ataxia type 7 (SCA7), a neurodegenerative disease caused by a polyglutamine expansion in the ataxin7 protein, in drosophila in which the expression of a mutant form of ataxin7 can be induced in neurons in adult flies without the confounding effects on development and on nonneuronal tissues. In this model, mutant ataxin7 accumulates in neuronal intranuclear inclusions containing ubiquitin, the 19S proteasome subunit and Hsp70, as in patients. Aggregation was accompanied by a decrease in locomotion and lifespan, but not by massive neuronal death. Disaggregation of the inclusions, when expression was stopped, correlated with improved locomotor function and increased lifespan, suggesting that the pathology may be susceptible to therapeutic intervention. Lifespan was then used as a quantitative marker in a genetic screen to identify several generic modulators of polyglutamine toxicity and specific modifiers of SCA7, involved in the normal function of ataxin7. This novel model is therefore a powerful tool for the study of SCA7 and other polyglutamine diseases and for the development of potential therapies. 961C Drosophila melanogaster as a model of mammalian hematopoiesis: characterization of the spaghetti mutant. Allyson Spence, Thomas Fellner, Margaret Fuller. Department of Developmental Biology, Stanford University School of Medicine, Stanford. Drosophila blood cells play important roles in immunity, wound healing, and development. Whereas mammals have a variety of terminally differentiated blood cell types of the myeloid and lymphoid lineages, the Drosophila blood system consists of only three terminally differentiated cell types. Plasmatocytes are similar to the mammalian monocyte lineage, and differentiate into phagocytic cells to encapsulate apoptotic cells and cell debris generated during stages of morphogenesis in the Drosophila life cycle. Innate imumunity and wound healing responses are the purview of the crystal cells. Lamellocytes are not observed in a healthy organsim, but rather undergo massive proliferation in response to specific immune challenges. Although the blood system of Drosophila is less complex than that of mammals, research over the past few years has revealed that mammalian and Drosophila hematopoiesis share remarkable similarities in their development and function. This makes Drosophila a viable model system to identify new genes involved in hematopoiesis. One setting which has been particularly fruitful for our understanding of mammalian blood cell development, is the study of melanotic tumors in Drosophila. Melanotic tumors represent over-proliferation of blood cells. Recently, several new mutants that cause melanotic tumors were identified in a genetic screen. In one intriguing mutant, spaghetti (spag), I observed that hematopoietic organs are increased in size and blood cells are elevated in number. Further analysis using specific markers showed that the phenotype is limited to cells of the crystal cell and lamellocyte lineage. As shown by homology searches, Spag has homology to members of the chaperone protein family. Indeed, preliminary data suggest that the mechanism of tumor formation is through its role as a chaperone, as treatment of larvae with geldanamycin, a specific inhibitor of the chaperone Hsp90, phenocopies the defect. Moreover, genetic data indicate that spag may be involved in the Toll signaling pathway through its interaction with Cactus, the Drosophila homolog of I-kB.

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47th Annual Drosophila Research Conference Late Poster Abstracts 962A Over-expression of JNK signaling rescues electromagnetic stress induced lethality in Drosophila. Kyu-Sun Lee1, Jang-Yul Kwak2, Dong-Seok Lee3, Kweon Yu1. 1) Development & Differentitation, KRIBB, Daejeon, KOREA; 2) Insect Resources Laboratory, KRIBB, Daejeon, KOREA; 3) Laboratory of Human Genomics, KRIBB, Daejeon, KOREA. Mobile phones are widely used in the modern world. However, biological effects of electromagnetic radiation from mobile phones are not well studied. In this report, we present biological effects of the 835MHz electromagnetic field (EMF), a mobile phone frequency, in the Drosophila model. Within the safety limit (specific absorption rate 1.6W/kg) from the American National Standard Institute, more than 90% of the flies were viable even after the 30hr exposure. However, in the strong EMF (specific absorption rate 4.0W/kg), the lethality was increased by the length of the EMF exposure. The strong exposure up-regulated stress response and apoptosis genes in the Drosophila cDNA chip analysis. This EMF exposure also increased the production of reactive oxygen species, induced apoptosis in the fly brain and whole body, and activated JNK signaling. When the gain-of-function JNK signaling mutants were expressed, the EMF induced lethality was significantly reduced. Over-expression of the anti-apoptotic capases inhibitor p35 significantly reduced the lethality, too. These findings demonstrate that the EMF exposure increased oxidative stress and induced apoptosis which may cause the increased lethality in Drosophila and this lethality was rescued by over-expression of JNK signaling or anti-apoptosis.

PHYSIOLOGY AND AGING 963B Regulation of SOD by LAMMER kinases. William Staatz, Sarah Wilkinson, Emmanuelle Meuillet. Arizona Cancer Ctr, Univ Arizona, Tucson, AZ. Accumulation of reactive oxygen species (ROS) in cells can lead to processes that promote disease states and cell death. Superoxide dismutase (SOD) and catalase (Cat) are the primary enzymes cells use to protect themselves from ROS. In the yeast. S. pombe, the genes for CuZn-SOD (sod1+) and Catalase (ctt1+) are upregulated by Lkh1+, a homolog of the LAMMER kinase family of serine threonine kinases (Cdc2-Like kinases). We examined the regulation of SOD and Cat by LAMMER kinases in Drosophila melanogaster and humans. The enzymatic activities of CuZn-SOD, Mn-SOD and Cat were measured in adult Drosophila mutants of Doa, the sole Drosophila LAMMER kinase, and in HeLa cells and MIAPaCa-2 human pancreatic carcinoma cells following treatment with the broad spectrum LAMMER kinase inhibitor, TG003. Our result show that in Doa mutants Mn-SOD activity is increased up to a 3.4-fold and CuZn-SOD activity is increased 2-fold. Treating HeLa and MIAPaCa-2 cells with TG003 resulted in a slight decrease in SOD2 by western blot and a doubling of CuZn-SOD activity in MIAPaCa-2 cells. Decreasing LAMMER kinase levels had no apparent effect on catalase activity in either flies or human cells. These data suggest that the regulation of SOD is a function of LAMMER kinases that has been broadly conserved during evolution. ____________________________

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x Future Drosophila Research Conferences x 2007 2008 Philadelphia, Pennsylvania Philadelphia Marriott Hotel San Diego, California Town and Country Resort & Convention Center March 711 April 26

47th Annual Drosophila Research Conference

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