04.

RNA splicing

75% dos genes humanos sofrem splicing

Modelo geral de splicing

DNA hnRNA

5 3

3 5

Nucleo
5

mRNA

Citoplasma

Tipos de processamento (splicing) -Nuclear (eucariotas) Sequencias consenso intro/exo (GT.....AG) snRNPs (small nuclear Ribonuclear Particles) U1.....U6 -Self splicing Mitocondria em algumas bacterias Grupo I -> Guanosina como factor (thermophila) Grupo II -> sequencia especfica (GT....AG) Intro/Exo (I e II possuem estruturas secundrias muito complexas) (RNA funciona como uma emzima Ribozyma) -Enzimatica ligase de splicing (S.cerevisiae, Xenopus)

O splisossoma utiliza os RNAs dos snRNPs para orientar a reao

Fungos filamentosos Mitocondria

Exon skipping or cassette exon: in this case, an exon may be spliced out of the primary transcript or retained. This is the most common mode in mammalian pre-mRNAs. Mutually exclusive exons: One of two exons is retained in mRNAs after splicing, but not both. Alternative donor site: An alternative 5' splice junction (donor site) is used, changing the 3' boundary of the upstream exon. Alternative acceptor site: An alternative 3' splice junction (acceptor site) is used, changing the 5' boundary of the downstream exon. Intron retention: A sequence may be spliced out as an intron or simply retained. This is distinguished from exon skipping because the retained sequence is not flanked by introns. If the retained intron is in the coding region, the intron must encode amino acids in frame with the neighboring exons, or a stop codon or a shift in the reading frame will cause the protein to be non-functional. This is the rarest mode in mammals.

Splicing Alternativo

Splicing occurs as transcription takes place

Alternative splicing: how to get more proteins out of your genome 5 DNA Nuclear Transcripts mRNAs Proteins 1 1 1 1 2 2 2 2 3 3 3 3 4 calcitonin 4 calcitonin 4 calcitonin 4 calcitonin
AAAA

5 CGRP

5 CGRP

Thyroid

5 CGRP

AAAA

Neurons

30,000 human genes probably give more than 100,000 different proteins

Down Syndrome Cell Adhesion Molecule

Axon guidance in the CNS

Alternative splicing of the -tropomyosin gene from rat. -tropomyosin is a coiled-coil protein that regulates contraction in muscle cells. The primary transcript can be spliced in different ways, as indicated in the figure, to produce distinct mRNAs, which then give rise to variant proteins. Some of the splicing patterns are specific for certain types of cells. For example, the -tropomyosin made in striated muscle is different from that made from the same gene in smooth muscle. The arrowheads in the top part of the figure demark the sites where cleavage and poly-A addition can occur.

Escolha do local de processamento

Splicing Alternativo (fast and slow Pol II)

RNA editing
Eucariotas Virus tRNAs rRNAs mRNAs miRNAs No RNA editing in prokaryotes

mRNA - Can alter AA sequence, splice site RNA degradation change from Adenosine to Ionosine leads to RNA degration RNA structure different protein binding site

RNA editing

Editing in trypanosomes

adenosine deaminases acting on RNA (Purine Biosynthesis)

Adenine:Inosine Inosine:citosine

Mechanism of A-to-I RNA editing in mammals. The position of an edit is signaled by RNA sequences carried on the same RNA molecule. Typically, a sequence complementary to the position of the edit is present in an intron, and the resulting double-stranded RNA attracts the A-to-I editing enzyme ADAR. Mice and humans have three ADAR enzymes: ADR1 is required in the liver for proper red blood cell development, ADR2 is required for proper brain development and the role of ADR3 is not yet known.

Apolipoprotein B