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Conservation and Evolution of Cis-Regulatory Systems
Tal El-HayComputational Biology Seminar
December 2005חנוכה תשס"ו
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""Nothing in biology makes sense except in Nothing in biology makes sense except in the light of evolutionthe light of evolution."."
- Theodosius Dobzhansky
Science, 23 December 2005
Evolution in Action
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Molecular Evolution
Wikipedia
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How Genomes Evolve
• Random mutations• Transposable DNA• Gene duplication• Divergence• Whole genome duplication (+divergence)• Recombination of exons
• Purifying selection
Molecular Biology of the Cell, Alberts et al., 4th ed.
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• “Relative dearth of species specific genes”• “Seeming abundance of functionally
homologous proteins”(A. P. Gasch et al., PLoS Biol., 2004)
Additional mechanisms for diversificationSuch as timing, location and level of
proteins
Our focus – Gene Expression
Evolution of Regulation
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Outline of Questions
• Conserved and evolved properties
• Mechanisms of conservation and evolution
• Bridging Genotype and Phenotype
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Cis-Regulatory Factors
• Composition
• Location
• Modules…
chiken A
mouse A
mouse 1
Gene control regions for eye lens chrystallins
Molecular Biology of the Cell, Alberts et al., 4th ed.
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Goal• Explore evolution of gene expression
regulation– Mainly through examination of cis-elements
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The Fungul Family
A. P. Gasch et al., PLoS Biol., 2004
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Large Scale Analysis
• Identify 264 co-regulated gene groups in S. serevisiae
• Putative cis-regulatory elements– 80 known consensus binding sites– 597 elements by motif search with MEME
• Score enrichment of genes containing each putative element- 42 cis-elements in 35 unique groups
• Orthologous modules in other species• Enrichment of orthologous modules
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Conservation of S. cerevisiae motifsG1 phase cell cycle ACGCG MCBAmino acid biosynthesis TGACTM Gcn4pNitrogen source GATAA GATA factors
Proteasome GGTGGCAAA Rpn4p
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Novel SequencesRibosomal proteins AGCCCTAARibosomal proteins GTGACTGTtRNA synthetases TGACTCAN
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Positions of binding sites
• Non random distribution• Similar across species• No correlations in locations
across species
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Spacing between binding sitesin Methionine Biosynthesis genes
• Small distance between Cbf1p and Met31/32p• Conserved across species• Independent of exact positions
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Zooming In – The Proteasome
• The proteasome is regulated by Rpn4p
• Consensus sequence enriched across all hemiascomycete
• Slight differences between C. Albicans and S. Serevisiae
Exploring evolution of the sequence
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Proteazome Cis-Element Evolution
Procedure:• Generate ‘meta matrix’
• Species specific matrix
• Hierarchical clustering
GGTGGCAAAW
AGTGGCAAAN
GGTGGCAAYA
GRAGGCAAAA
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Binding specificities of Rpn4p
Sc elementCommon elementCA element
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Validation of specificities?
Sc_Rpn4p
Ca_Rpn4p
hybrid
A. GGTGGCAAAAB. GAAGGCAAAAC. AGTGGCAACA
D. GGTGGCAAAAE. AAGTGGCAAAAF. GGTGGCAACCAG. CTGCATTTGG
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Intermediate Summary - Conservation
• Conserved cis-elements in ‘un-align-able’ non coding regions
• Correspondence of conservation and evolutionary distance
• Did not observe position conservation
• Similar position distributions
• Found an example of conserved spacing– Interaction constraint between TF?
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Intermediate Summary - Evolution
• Novel sequences in coregulated orthologous modules
• Conserved expression patterns with evolved regulation (e.g. Proteasome, Ribosome)
• Individual example for:– Addition of gene targets to a regulatory
network (S1 phase cell cycle)– Coevolution in a regulatory network– Cooption of a regulatory system
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Goals• Learn mechanisms of evolution of regulation
systems• Integrating comparative expression and
sequence analysis
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Computational framework
Identify conserved modules
Derive orthologous modules
Identify cis-elements profiles
Reconstruction of evolution
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Orthologous Modules in Yeast
a. S-phase module
b. Respiration
c. Amino acid metabolism
d. Ribosomal proteins synthesis
e. Stress
f. Ribosome biogenesis
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Conserved and Diverged Regulatory Mechanisms
• Conserved cis-elements – Respiration module – Mlu cell cycle element (ACGCGT)– Bound by MBF complexes in both species
• Diverged - Ribosomal protein synthesis– RAP1 and IFHL in S. cerevisae
(TACATCCGTACAT & TCCGCCTAG resp.)– Homol-D box and Homol-E site in S. pombe
(TGTGACTG & ACCCTACCCTA)
• Conserved and diverged – Ribosome biogenesis module
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The evolution of the Ribosomal Regulatory Program
Apparent redundancy ofbinding sites
Switching from Homol-Dto RAP1
Gradual Evolution in the IFHL Box
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Evidence for Regulator Switching
• Evolution of Transcription factor– RAP1 is a submotif of telomeric
repeat– Rap1p regulates telomer length– Association of events:
Rap1p Gained of Trans activation domain, RAP1 joined RP module
• Interacation of RAP1 & Homol-D?– usually 2-6 base pairs apart– Conserved order
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Gradual Evolution of IFHL Box
TCTGCCTA
AGGGCTATAGCCCT
GCCCTA
CCCTACCCTA
Convergent domain duplicationor
Acquired dimerization domain?
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Spatial Configuration• HomolD-RAP1
Sites are tightly coupled
• Rap1-IFHLVariable probably to modified role of IFHL
• RAP1-RAP1Sites are tightly coupled
• IFHL-IFHLCoupling mainly inA. gossypii and K. Waltii
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Intermediate Summary -Modes of Evolution
• Conserved modules with diverged regulatory mechanisms
• Some changes via redundant intermediate programs
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Goals
• Genotype and Phenotype
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Background – Yeast Growth
• Some Yeast species are fermentative
• Others can employ only respiration
• Connection to whole genome duplication
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Analysis of Genetic Basis of Phenotypic Diversity
• Examine gene expression program– S. Cerevisiae with 1000 expression profiles – C. Albicans with 198 profiles
• Motif search in related modules
• Validation of motif role
• Comparison of motif and phenotype evolution
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Transcriptional Wiring Differences
RP Cytoplasmic ribosomal proteinsrRNA rRNA processing genesMRP Mitochondrial RPSTR Environmental Stress response
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Cis or Trans?
Cis element search:• PAC in rRNA of both
species• None for MRP in
S. Serevisiae• AATTTT in MRP,
putative rRNA regulator
Validation of cis-regulatoryRole of AATTTT in MRP
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Spatial Configuration of AATTTT
Both species:• Position is confined in RP
and rRNA• Not represented in STR
C. Albicans:• AATTTT Regulates also
MRP
Rapid growth element
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Loss or Gain of Binding Site?
• Loss of binding site in MRP associated with whole genome duplication
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Evolution of Genome and Phenotype
• “Gene duplication can facilitate the evolution of new function”– by specialization of new coding sequences– Also by facilitating the evolution of gene
expression
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Summary of methodologies
• Integration of sequence and gene-expression– Finding orthologous modules– Finding orthologous binding motifs
• Exploiting phylogenetic trees– Find genotype change rules– Associate phenotype and genotype changes– Exploit gene expression data of extremes
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Summary of principles
• Conservation of regulatory programs– Binding site conservation– Position and spacing
• Conservation of module with evolution of control– Loss and gain– Drift– Switching, could be explained via redundancy
• Evolution of regulatory programs -> Evolution of phenotype– Addition of gene targets– Cooption or loss of TF– Also facilitated by whole genome duplication
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Points for thought
• How can simultaneous change of cis-elements happen?
• Evolution of elaborate wiring
• Evolution of other modes of regulation
• Is there gene expression data for more species?
• Look for conserved patterns?