Journal Club 2009

Science. 2009 April 17;324(5925):389-92

Local DNA Topography Correlates with
Functional Noncoding Regions of the Human
Genome
Stephen C. J. Parker, Loren Hansen, Hatice Ozel Abaan, Thomas D. Tullius, Elliott H.
Margulies

João Carneiro
jcarneiro@ipatimup.pt
Is structural information important when analysing the DNA molecule?

Human genome

≈ 2% code for ≈ 98% non-coding

proteins Regions not well
understood
Functional Association to conserved regions
regions because of their importance to
the organism

The analysis is based in the

primary nucleotide sequence

Evolutionary sequence-constraint
algorithms
DNA is a double helix molecule
with a three dimensional
structure that varies according to
the nucleotide sequence.

Shape of
grooves

Shape of backbone
 Previous described method to predict the modification
of the DNA three dimensional structure

Hydroxyl radical cleavage

Substitutions of a Similar Very different Similar sequences

few bases can sequences sequences at can adopt very
affect the often adopt primary level can different
cleavage pattern similar have identical structures (few)
structures structure
Hydroxyl radical
footprinting of RNA

Gel pattern of
unfolded RNA is
cleaved uniformly

Gel pattern of
folded RNA is
cleaved where the
solvent has
accessibility to the
backbone
High similarity in cleavage patterns for two sequences with low
sequence identity. Plotted
are the hydroxyl radical cleavage patterns of two 10-mer sequences that
share no common nucleotides (sequence identity = 0%). Note the
significant correlation (R = 0.94) of the two patterns.
Construction of a genome-scale structural map at single-
nucleotide resolution
Jason A. Greenbaum, Bo Pang and Thomas D. Tullius
Genome Res. 2007 17: 947-953
 ORChID database

Collection of a library of hydroxyl radical cleavage

experimentally determined for 4-bp DNA sequences

Prediction of the DNA backbone and grooves of genomic DNA

in all possible 11–bp sequences

In silico library of structural

profile of DNA

Euclidean distance to detected conserved structure

possible represents important biological functions
Quantification of single base substitutions in DNA structure
The distribution of average structure changes observed for all
11-bp sequences
Alignments of 30 Mb of high quality comparative sequence data for 36
different species

Chai: Structural information + DNA sequence–based binomial

conservation (binCons)
binCons: only DNA primary sequence analised

Incorporation of a false discovery rate (FDR)

Random permutation of alignment columns to
generate a new alignment (total number of
constrained bases are compared with the original
alignment and confidence value determined)

Re-run of Chai and binCons algorithm over null alignments

For all statistical confidence values Chai identified more constrained
region than binCons
 Identification of the regions retrieved by Chai that harbor functional
elements

DNase I Predicted transcriptional Ancestral repeats

hypersensitive enhancers identified
sites using chromatin
modification patterns

Structural
characteristics and
nucleotide sequence
are important in non-
coding regions
Analysis of nucleotide
substitutions consequences in
structure profile of DNA binding
proteins

Mammal transcription factor

Zif268 protein and archaeal
transcriptional regulator Ss-
LrpB

Results revealed that low-

affinity binding sites
differed dramatically in
structure from high-affinity
sites

DNA structural profiles correlate with motif-

binding affinity for the mutant REDV Zif268
protein.
Collection of 734 noncoding single-nucleotide variants in the
human genome
associated with a phenotype (PhenCode Project)

For comparison, a distribution of baseline variation in DNA

topography was computed for 16,832 neutrally evolving
single-nucleotide polymorphisms (SNPs)

Analyses of 11-bp window centered on the nucleotide variant

using Chai
Big alterations in DNA structures are significantly correlated to non-
coding phenotypic associated variants

P < 3 × 10−4
Wilcoxon rank sum test

Distribution of DNA structural changes for phenotype associated variants (red

line) and neutral variants (black line)
 Finally they analysed 12 predicted enhancer-
containing regions

7 regions overlap a 5 regions overlap

combination of Chai- elements detected
and binCons-detected only by the Chai
elements algorithm

They used a luciferase reporter construct that

included this regions and transfected them into 293T
cells
 Luciferase-based reporter activity of 12 regions
containing Chai detected elements

P ≤ 0.05
Wilcoxon rank sum test
 Conclusions

DNA local structural architecture (DNA topography) is wide

conserved in the analysed regions

These regions overlap the majority of known non-coding

functional sites

Conserved regions that escaped detection by DNA primary

sequence analyses can be detected by this method