Clustered cis-regulatory elements underlie adaptive divergence in sticklebacks Felicity Jones Friedrich Miescher Laboratory of Max Planck Society Tübingen, Germany Male threespine stickleback
Molecular mechanisms of adaptation & speciation 1 1 The regulatory control of adaptive divergence in gene expression 2 Functional dissection of adaptive regulatory elements 3 The evolution & role of recombination during adaptive divergence
Threespine sticklebacks have undergone a recent adaptive radiation Bell & Foster (1994) Extensive morphological, physiological, behavioural diversity Marine Marine Freshwater eshwater
Repeated & independent evolution provides biological replicates of the evolutionary process Marine Marine Freshwater eshwater Benthic Benthic Limnetic Limnetic
Whole genome sequencing of marine–freshwater species pairs revealed parallel adaptive divergence at ~81 predominantly intergenic genomic loci High resolution map of 81 parallel marine-freshwater adaptive loci (FDR 0.02) Jones et al (2012) Nature
Suggests two molecular mechanisms play a role in parallel adaptive divergence of sticklebacks: 1 1 Regulatory mechanisms Jones et al (2012) Nature
Suggests two molecular mechanisms play a role in parallel adaptive divergence of sticklebacks: How is adaptive divergence in gene expression regulated? 1 1 Regulatory mechanisms cis mRNA What are the mutations & functional elements? Jones et al (2012) Nature
Suggests two molecular mechanisms play a role in parallel adaptive divergence of sticklebacks: Adaptive loci (marine-freshwater divergence) 0.5cM/Mb The genomic recombination landscape 2 shapes the loci of adaptation Jones et al (2012) Nature
How is adaptive divergence in gene expression regulated? Identifying adaptive gene expression differences using comparative transcriptomics Dr Jukka-Pekka Marine Freshwater eshwater Verta Verta & Jones (in review) BioRxiv doi: https://doi.org/10.1101/412932
How is adaptive divergence in gene expression regulated? Identifying adaptive gene expression differences using comparative transcriptomics Dr Jukka-Pekka Marine Freshwater eshwater Verta RNAseq RNAseq of of Gill Gill Tissue Tissue Freshwater eshwater Expr Expression ession (RPKM) (RPKM) Marine Marine Expression Expr ession (RPKM) (RPKM) Genes Genes Verta & Jones (in review) BioRxiv doi: https://doi.org/10.1101/412932
How is adaptive divergence in gene expression regulated? Identifying adaptive gene expression differences using comparative transcriptomics Dr Jukka-Pekka Marine Freshwater eshwater Verta RNAseq RNAseq of of Gill Gill Tissue Tissue Freshwater eshwater Expression Expr ession (RPKM) (RPKM) Marine Marine Expression Expr ession (RPKM) (RPKM) Genes Genes Adaptive gene expression differences (parallel divergence in gene expression) Verta & Jones (in review) Freshwater eshwater-lik -like e Marine-lik Marine-like e Composite PC Composite PC BioRxiv doi: https://doi.org/10.1101/412932
Molecular signatures of natural selection acting on genes with parallel gene expression divergence Elevated genetic divergence (F ST MAR v FW ) around TSS loci with parallel expression divergence Marine-Freshwater control loci Reduced nucleotide diversity (pi) around TSS Marine Freshwater control loci control loci loci with parallel expression divergence Verta & Jones (in review) loci with parallel expression divergence BioRxiv doi: https://doi.org/10.1101/412932
How are adaptive expression differences controlled? trans vs cis- regulation cis mRNA Marine Freshwater eshwater trans F1 Hybrids F1 Hybrids Verta & Jones (in review) BioRxiv doi: https://doi.org/10.1101/412932
Allele-Specific Expression assays: RNAseq of F1 hybrids to determine cis- vs trans- regulation Allele specific expression in F1 Hybrid (log freshwater allele /marine allele) Genes regulated in trans Parental Expression Divergence (log freshwater/marine) Verta & Jones (in review) BioRxiv doi: https://doi.org/10.1101/412932
Cis-regulation is the predominant mechanism underlying gene expression divergence Genes regulated in cis Genes regulated in trans Verta & Jones (in review) BioRxiv doi: https://doi.org/10.1101/412932
Cis-regulation is the predominant in multiple mar-fw pairs (and is associated with the degree of genome-wide genetic divergence) Marine Freshwater eshwater Proportion of Transcripts 0.25 Tyne yne Forss orss Shiel Shiel Little C Lit tle Campbell ampbell 1.62M 1.54M 2.27M 2.75M 0.20 0.15 0.10 0.05 0.00 cis cis cis cis cis cis cis cis tr trans cis+tr cis+trans cis cis-tr compensatory compensatory tr trans cis+tr cis+trans cis-tr cis compensatory compensatory tr trans cis+trans cis+tr cis cis-tr compensatory compensatory trans tr cis+trans cis+tr cis cis-tr compensatory compensatory ans ans ans ans -trans -trans -trans -trans ans ans ans ans ans ans ans ans Verta & Jones (in review) BioRxiv doi: https://doi.org/10.1101/412932
Gene regulatory mechanisms and their magnitude evolve in parallel across populations Correlation in cis-regulation � among populations � (Pearson’s r) � Genes upregulated � Genes with � in freshwater � no difference � relative to marine � in expression � Degree of Parallel Divergence in Expression � Verta & Jones (in review) BioRxiv doi: https://doi.org/10.1101/412932
The degree of parallelism in gene regulation appears to be associated with regulatory effect size Correlation in cis-regulation � among populations � (Pearson’s r) � Genes upregulated � Genes with � Genes downregulated � in freshwater � no difference � in freshwater � relative to marine � in expression � relative to marine � Verta & Jones Degree of Parallel Divergence in Expression � (in review) BioRxiv doi: https://doi.org/ 10.1101/412932
Cis- regulation is more additive than other forms of gene expression regulation – more visible to natural selection Dominant Dominant Additive Verta & Jones (in review) BioRxiv doi: https://doi.org/ Dominance : Additivity 10.1101/412932
Cis- regulation is more additive than other forms of gene expression regulation – more visible to natural selection Dominant Dominant Additive And stable under different environmental conditions Siblings raised under different environmental conditions show strong correlation in cis-regulation Verta & Jones (in review) BioRxiv Correlation in allele-specific doi: https://doi.org/ expression Dominance : Additivity 10.1101/412932
Cis- regulation is more additive than other forms of gene expression regulation – more visible to natural selection Dominant Dominant Additive And stable under different environmental conditions Siblings raised under different environmental conditions show strong correlation in cis-regulation Correlation in gene expression Verta & Jones (in review) BioRxiv Correlation in allele-specific doi: https://doi.org/ expression Dominance : Additivity 10.1101/412932
Molecular mechanisms playing a role in parallel adaptive divergence of sticklebacks: How is adaptive divergence in gene expression regulated? 1 1 Regulatory mechanisms cis mRNA What are the mutations & functional elements? Jones et al (2012) Nature
High resolution signals, good for functional follow-up … but ancient haplotypes carry many candidate mutations EDA Marine – Freshwater CSS MAR - FW Divergence CSS 20kb 40kb <5kb Genes Freshwater Raw Data Jones et al (2012) Nature Fr Ra Marine Marine
Identifying regulatory elements Dr Stanley Neufeld underlying divergent adaptation Comparative functional annotations of adaptive loci using epigenomics ChipSeq, ATACseq, RNAseq & CaptureC Mapping poised & active enhancers, open chromatin differential expression and regulatory interactions in marine and freshwater genomes
Open chromatin and epigenetic modifications to histones are predictive of functional and active regulatory elements Modified from image from ENCODE - NHGRI & EBI
HMM HMM epigenet epigenetic states ic states Histone Histone modificat mod ifications: ions: Marine males H3K27ac ChIPseq Marine females H3K4me1 ChIPseq Freshwater males H3K4me3 ChIPseq Freshwater females F1 Hybrids Outgroups Open chromat Open chr omatin in ATACseq Liver Kidney Gill Gene expression Gene expr ession RNAseq Transcript ranscript annotat annotations ions
Marine v Fr Marine v Fresh esh diver ivergent gent HMM HMM enhancers enhancers epigenet epigenetic states ic states Histone Histone mod modificat ifications: ions: Marine males H3K27ac ChIPseq Marine females H3K4me1 ChIPseq Freshwater males H3K4me3 ChIPseq Freshwater females F1 Hybrids Outgroups Open chr Open chromat omatin in ATACseq Liver Kidney Gill Marine v Fr Marine v Fresh esh diver ivergent gene gent gene expression expr ession Gene expression Gene expr ession RNAseq Transcript ranscript annotat annotations ions
Enhancer marks are enriched around Transcription Start Sites Chromatin State open chromatin H3K27ac H3K27ac & H3K4me1 H3K4me1 no mark Freshwater open chromatin H3K27ac H3K27ac & H3K4me1 H3K4me1 no mark Marine Fold enrichment 0 5 10 15
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