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2/3/2015 Sex chromosomes demonstrate complex evolutionary trajectories across bird taxa Sex is often determined by a pair of heteromorphic sex chromosomes. Doris Bachtrog University of California, Berkeley I. Introduction Sex chromosomes evolve from autosomes autosomes XY X X X X X X X X X X X X XY XY Sry Sry X X X I. Introduction Same evolutionary steps in ZW systems XY Sry Sry X autosomes Z W X X X X X X X X X X X X ZW ZW DMRT1 DMRT1 X X X ZW DMRT1 DMRT1 X - 1st step: Genetic sex determination - Recombination suppression evolves in independent steps (evolutionary strata) - X & Y differentiate; older evolutionary strata are more diverged - 1st step: Genetic sex determination - Recombination suppression evolves in independent steps (evolutionary strata) - Z & W differentiate; older evolutionary strata are more diverged - Y chromosomes degenerate and may accumulate male-specific genes - X chromosomes often become dosage compensated and may evolve specialized gene content (demasculinization) - W chromosomes degenerate and may accumulate female-specific genes - Z chromosomes evolve specialized gene content (masculinization) but no dosage compensation has been found yet in ZW systems I. Introduction Sex chromosomes are at different stages of differentiation in different lineages XY XY X X X X X X X X X homomorphic sex chromosomes X X X X ZW XY Sex chromosomes are at different stages of differentiation in different strata ZW X X X X X I. Introduction X heteromorphic sex chromosomes XY XY X X X X X X X X X homomorphic sex chromosomes X X X X ZW XY ZW X X X X X X heteromorphic sex chromosomes 1 2/3/2015 II. Bird sex chromosome evolution Sex chromosome evolution in birds Sex chromosomes in birds autosomes PALEOGNATHAE X X X X X X X Z W ZW ZW X X X X X X X ZW DMRT1 DMRT1 DMRT1 DMRT1 X ? - All birds have homologous sex chromosomes NEOGNATHAE - DMRT1 appears to be the master sex determining locus - In emu/ostrich: large PAR’s - In chicken: W fully degenerated, evolutionary strata; no dosage compensation of Z I. Introduction I. Introduction Using male/female coverage to determine X-derived scaffolds Identify sex-linked genes using genomic sequencing Female and Male PE reads Scaffolds I. De novo Assembly DNA II. Map reads to scaffolds & determine coverage DNA III. Map scaffolds to reference genome & plot coverage 1 - de novo assembly of genome 2 – determine sex-linkage using male and female coverage 0.5 PAR DR PAR Coverage Coverage Reference genome 1 Autosomal I. Introduction II. Bird sex chromosome evolution Using male/female coverage to determine Y-derived scaffolds Female and Male PE reads Sex chromosomes in birds Scaffolds I. De novo Assembly Paleognathae Galloanserae II. Map reads to scaffolds & determine coverage In collaboration with BGI: Genome sequences of 18 bird species from females (ZW system) High-quality assemblies from ostrich and chicken; other genomes were scaffolded against ostrich genome (for Paleognathae) or against chicken (for Neognathae) Neoaves III. Map scaffolds to reference genome & plot coverage X-linked Use coverage from females only to identify sex-linked scaffolds young stratum IV. Y-linked scaffolds may show some homology with X chromosome Coverage old stratum Coverage Reference genome Y-linked X-linked Neognathae 2 2/3/2015 II. Bird sex chromosome evolution Using male/female coverage to determine Z-derived scaffolds Female and Male PE reads Scaffolds I. De novo Assembly PAR’s II. Map reads to scaffolds & determine coverage III. Map scaffolds to chicken / ostrich Z & plot coverage 0.5 DR PAR PAR (Pseudo)Autosomal Z-linked II. Bird sex chromosome evolution Using male/female coverage to determine W-derived scaffolds Female and Male PE reads Ostrich Z Size of PARs varies dramatically across lineages – independent evolution of sex chromosomes in different bird lineages To reconstruct the evolutionary history of sex chromosomes in birds: Identify candidate Wregions as un-mapped scaffolds with Z homology Coverage 1 Coverage Chicken/ostrich Z DMRT1 II. Bird sex chromosome evolution Coverage analysis along Z reveals dramatic differences in PAR among birds Chicken Z II. Bird sex chromosome evolution W-linked fragments in birds Scaffolds I. De novo Assembly II. Map reads to scaffolds & determine coverage III. Map scaffolds to chicken / ostrich Z young stratum IV. Infer strata based on sequence homology between Z & W fragments Coverage old Chicken/ostrich Z Coverage DMRT1 W-linked Z-linked II. Bird sex chromosome evolution W-linked fragments in birds II. Bird sex chromosome evolution W-linked fragments reveal evolutionary strata W-fragments differ in density & ZW similarity -> strata Allow us to infer the evolutionary history of bird sex chromosomes: Older strata: less homology between Z & W Not fully degenerate W’s 3 2/3/2015 II. Bird sex chromosome evolution Are W-candidates indeed W-linked? Exact same mapping pipeline applied to chromosome 1: II. Bird sex chromosome evolution Are W-candidates indeed W-linked? Coverage of W-candidates, Z-linked sequences and autosomes in males and females W-sequences are femalelimited Z-sequences show half the coverage in females Autosomal sequences show similar coverage in males and females - No differentiated region on chromosome 1 - Very little mapping of unmapped fragments II. Bird sex chromosome evolution Evolutionary strata of bird sex chromosomes Age of strata and gene synteny information allow us to reconstruct evolutionary history of bird sex chromosomes II. Bird sex chromosome evolution Evolutionary strata of Paleognathae Shared and lineage-specific strata in bird sex chromosomes S0 is shared among all birds - High overlap of gene content Gametologs cluster by chromosome and not species ZW divergence time exceeds species divergence time Other strata in Palaeognaths are lineage-specific - - DMRT1 is in ancestral stratum in each species Little overlap of gene content Gametologs cluster by species and not chromosome ZW divergence time is less than species divergence time - Strata become progressively younger when moving from ancestral stratum to PAR II. Bird sex chromosome evolution Evolutionary strata of Neognathae II. Bird sex chromosome evolution Use synteny to infer if inversion contribute to strata formation Shared and lineage-specific strata in bird sex chromosomes S1 is shared among all Neognathae S2 is shared among all Neoaves but not Galloanserae S3 is PAR / lineage-specific 4 2/3/2015 II. Bird sex chromosome evolution Sex chromosome differentiation in birds II. Bird sex chromosome evolution Sex chromosome differentiation in birds II. Bird sex chromosome evolution Sex chromosome differentiation in birds II. Bird sex chromosome evolution Sex chromosome differentiation in birds II. Bird sex chromosome evolution Sex chromosome differentiation in birds II. Bird sex chromosome evolution Sex chromosomes of birds Shared sex chromosomes but lineagespecific events have generated a diversity of sex chromosome karyotypes! 5 2/3/2015 II. Bird sex chromosome evolution II. Bird sex chromosome evolution W degeneration over time W degeneration over time How many genes are left on W chromosomes? Younger strata contain more intact genes White-throated Tinamou Z W 2 strata with W-linked genes II. Bird sex chromosome evolution II. Bird sex chromosome evolution W degeneration over time Conclusions Avian sex chromosomes harbor diverse compositions of DR and PAR across taxa Avian W chromosomes showed great variation in their degree of degeneration the putative avian sex determining gene DMRT1 is consistently within the nonrecombining region of the sex chromosomes avian sex chromosomes have suppressed recombination through a series of punctuated events (“strata”) with a gradient of ages, reflected by higher levels of sequence homology between W and Z sequences as one moves towards the PAR and away from DMRT1 reconstruct the evolutionary history of recombination suppression and differentiation across birds dynamics of W degeneration changes over time, with W-linked genes subject to ongoing decay, and the tempo of W degeneration slowing down in older strata Rate of gene loss is not linear over time (but declines exponentially) III. Snakes Sex chromosomes of snakes Sex chromosome evolution in snakes - Species with homomorphic and heteromorphic sex chromosomes & some in between autosomes Z W X X X X X X X X X X X X ZW ? ZW X X X ZW ? ? ? X boas & pythons colubridae vipers & cobras Closely related species (i.e. garter snakes; Colubridae) appear to have both homomorphic and heteromorphic sex chromosomes Rapid evolution of sex chromosomes???? - All snakes have homologous ZW sex chromosomes - In boas & pythons: largely undifferentiated sex chromosomes - In Colubridae: various intermediate stages of differentiation which suggests that ancestor of Colubrids had homomorphic sex chromosomes and many speciesspecific events created diverse sex chromosomes in Colubrids - In vipers & cobras: fully degenerate W chromosomes; dosage compensation status unknown 6 2/3/2015 III. Snakes III. Snakes Anolis genome to identify snake sex chromosomes No dosage compensation in snakes Anolis chromosome 6 corresponds to snake sex chromosome Less expression from Z in female rattlesnakes – no dosage compensation! Boa rattlesnake - Boa has completely homomorphic sex chromosomes - Sex chromosomes of rattlesnake and garter snake are completely differentiated III. Snakes Evolutionary strata on snake sex chromosomes Evolutionary strata on snake sex chromosomes No homology between Z and W at middle region – central part stared to differentiate first! garter snake 2 or 3 evolutionary strata on snake Z chromosome! rattlesnake III. Snakes x x x x x x x x x no differentiation in boas x x x x x x x x cytogenetic data gave wrong impression of variation among sex chromosomes in Colubridae x x x x x x evidence for 2 shared strata in garter snakes & rattle snakes IV. Flies Basic karyotype in Diptera Anopheles In many Diptera families: 6 pairs of chromosomes (5 rods, 1 dot) IV. Flies Basic karyotype in Diptera Sex chromosome in Drosophila & Anopheles In many Diptera families: Element A Anopheles 6 pairs of chromosomes (5 rods, 1 dot) Element B Element C Element D Element E Gene content among chromosomes conserved Gene content among chromosomes conserved Muller element’s Muller element’s Drosophila Element F Drosophila 7 2/3/2015 IV. Flies IV. Flies Sex chromosome evolution across Diptera crane flies Coboldia fuscipes dung midges Maye ola destructor Herme a illucens Condylostylus pa bulatus Megaselia abdita Eristalis dimidiata Themira minor chironimids black flies mosquitoes sand flies Tipula oleracea Trichoceridae sp. crane flies Coboldia fuscipes dung midges gall midges Maye ola destructor gall midges solder e fli s robber flies long-legged flie s Herme a illucens solder e fli s robber flies long-legged flie s de novo gain of sex chromosomes in some lineages hump-back flies hover fli s scavenger flies Megaselia abdita hump-back flies hover fli s scavenger flies some species have entirely lost sex chromosomes Holcocephala fusca Condylostylus pa bulatus Eristalis dimidiata Themira minor true frui lies Eutreta diana Tephri s californica (ZW) Trupanea negronis Bactrocera oleae (XY) true frui lies Teleopsis dalmanni Sphyracephala brevicornis stalk-eyed flies Teleopsis dalmanni Sphyracephala brevicornis stalk-eyed flies Liriomyza trifolii leaf miners Liriomyza trifolii leaf miners Ephydra hians Ephydra e gracilis Phor ca variegata shoree fli s Ephydra hians Ephydra gracilis Phor ca variegata shoree fli s Drosophila pseudoobscura Drosophila miranda Drosophila melanogaster Drosophila busckii Drosophila albomicans BRACHYCERA Eutreta diana Tephri s californica (ZW) Trupanea negronis Bactrocera oleae (XY) Drosophila rela ves Sex-linked e element(s) A B C D E F Scaptodrosophila lebanonensis Glossina morsitans tsetse flies Sarcophaga bullata Sarcophagidae sp. Lucilia sericata Calliphora erythrocephala flesh fli s bot flies bloweflies Undifferen ated Vicoso & Bachtrog, PLoS Biology, in press Drosophila pseudoobscura Drosophila miranda Drosophila melanogaster Drosophila busckii Drosophila albomicans Drosophila rela ves Scaptodrosophila lebanonensis Glossina morsitans tsetse flies Sarcophaga bullata Sarcophagidae sp. Lucilia sericata Calliphora erythrocephala flesh fli s bot flies bloweflies a dozen different sex chromosome karyotypes NEMATOCERA Tipula oleracea Trichoceridae sp. Holcocephala fusca Chironomus riparius Chaoborus trivi atus Mochlonyx cinc pes Anopheles gambiae Aedes aegyp Clogmia albipunctata >30 species, 20 different families >250 MY evolution ancestral karyotype: dot chromosome as sex chromosome or homomorphic sex chromosomes some families have replaced sex chromosomes with different Muller element BRACHYCERA chironimids black flies mosquitoes sand flies NEMATOCERA Chironomus riparius Chaoborus trivi atus Mochlonyx cinc pes Anopheles gambiae Aedes aegyp Clogmia albipunctata Sex chromosome evolution across Diptera some families have incorporated additional chromosomes into their sex chromosomes Muller element D is repeatedly used as a sex chromosome XY to ZW transition in Tephritidae A B C D E F Undiff IV. Flies Conclusions X chromosomes show partial dosage compensation Transcriptome analysis of whole flies to study functional properties of sex chromosomes P. variegata T. minor * *** E. hians L. trifolii *** * 2 0.5 2 4 2 Sex-chromosome turnover has occurred multiple times throughout Dipteran evolution; bird & snake sex chromosomes are conserved but show signs of evolutionary strata -1 -3 -4 -1.0 -6 -1.5 -2 -2 -1.0 -2 -4 -2 -0.5 -1 -2 -0.5 0 0 0 0 0.0 0 0.0 1 2 0.5 1 2 0 Log2[M/F expression] -2 -4 X X=A X X=B A A A A X X=C,D,F Comparing male to female coverage can be a good approach to identify sexlinked sequences in non-model organisms, and combined with gene expression analysis, this allows us to make general inferences on sex chromosomes biology 1.0 2 3 6 G. morsitans 4 M. destructor 1.0 *** 1.5 T. dalmanni *** 4 D. melanogaster AA X X=A A A X X=A,D,F AA X X=D A A X X=D A A X X=D A A Dosage compensation has evolved in multiple Diptera families, through upregulation of the X in males; dosage compensation is lacking in snakes & birds This provides a great sample in which to test hypothesis of sex-chromosome evolution in a more systematic manner than has so far been possible. Expression of single X in males in not half that of females! Always some level of dosage compensation, but perhaps some evidence of specialized gene content as well. Acknowledgements UC Berkeley Beatriz Vicoso Qi Zhou Chris Ellison Wynn Meyers Karen Wong Tania Gurbich Emily Brown Lauren Giblisco Shivani Mahajan BGI Jilin Zhang Guojie Zhang Zaak Walton 8