Y chromosome dynamics in Drosophila. Amanda Larracuente Department of Biology

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1 Y chromosome dynamics in Drosophila Amanda Larracuente Department of Biology

2 Sex chromosomes J. Graves X X X Y

3 Sex chromosome evolution Autosomes Proto-sex chromosomes Sex determining Suppressed recombination Differentiation X Y Reviewed in Rice 1996, Charlesworth 1996

4 Y chromosomes Male-restricted Non-recombining Degenerate Heterochromatic Image from Willard 2003

5 Drosophila Y chromosome D. melanogaster Cen ~20 genes ~40 Mb Hoskins et al Acquired from autosomes Heterochromatic: Ø 80% is simple satellite DNA Photo: A. Karwath Lohe et al. 1993

6 Satellite DNA Tandem repeats Heterochromatin Centromeres, telomeres, Y chromosomes Yunis and Yasmineh

7 Y chromosome assembly challenges Repeats are difficult to sequence Underrepresented Difficult to assemble Genome Sequence read: Short read lengths cannot span repeats

8 Single molecule real-time sequencing Pacific Biosciences Average read length ~15 kb Long reads span repeats Better genome assemblies Zero mode waveguide Eid et al. 2009

9 Comparative Y chromosome evolution in Drosophila I. Y chromosome assemblies II. Evolution of Y-linked genes

10 Drosophila genomes 2 Mya 0.24 Mya Photo: A. Karwath P6C4 ~115X ~120X ~85X ~95X

11 De novo genome assembly Assemble genome Iterative assembly: Canu, Hybrid, Quickmerge Polish reference Quiver x 2; Pilon 2L 2R 3L 3R 4 X Y Assembled genome Mahul Chakraborty, Ching-Ho Chang

12 De novo genome assembly species Total bp # contigs NG50 D. simulans 154,317, , D. mauritiana 154,866, ,121,759 D. sechellia 166,750, ,907,079 2L 2R 3L 3R 4 X Y Assembled genome Mahul Chakraborty, Ching-Ho Chang

13 Genome assembly Missing most of Y chromosome! 2L 2R 3L 3R 4 X Y Assembled genome

14 Underrepresented heterochromatin Sequenced males Expected: ~100X for autosomes ~ 50X for X ~ 50X for Y count Dmau Y X Region A U X Y Observed: ~100X for autosomes ~ 50X for X ~ 35X for Y Ching-Ho Chang coverage coverage

15 Y chromosome assembly Map reads to reference genome Remove X and autosomal reads 2L 2R 3L 3R 4 X Y

16 Y chromosome assembly Map reads to reference genome Remove X and autosomal reads Reassemble leftover reads Manual curation Confirm Y contigs Y X/A heterochromatin

17 3000 Identifying Y-linked contigs A Location count Illumina reads Unknown 1000 Map XX Map XY Take ratio count Y X X Location Y Unknown X Y Female/Male Female/Male

18 Y-sensitive assemblies X + autosomes Reference + Y enriched merge New reference 2L 2R 3L 3R 4 X Y

19 Y chromosome assembly ~40 Mb Assembly con,gs total size N50 Dmel reference (v6 ) 261 3,977,036 81,922 Dmel PacBio 97 15,245, ,400 Dsim PacBio 40 14,132,328 1,189,645 Dsec PacBio 68 14,889, ,900 Dmau PacBio 19 14,537,038 2,282, % of Y chromosomes in contigs! Ching-Ho Chang

20 Dmel Y-linked genes Most Y-linked genes acquired from autosomes 2L 2R 3L 3R 4 X Y e.g. Carvalho et al. 2000, 2001; Carvalho et al. 2015; Krsticevic et al. 2015

21 Dmel Y-linked genes origin Dmel Dsim Dsec Dmau kl-5 3R PRY 2L kl-3 2L kl-2 2R Pp1-Y1 2R ARY 3L Ppr-Y 2L WDY 2L Pp1-Y2 3R ORY 3R CCY 3R Mst77Y1-18ψ 3L FDY 3R Mst35Y 2R Consistent with Koerich et al. 2008; labs of AB Carvalho and AG Clark

22 Y-linked gene organization Dmel cen kl-5 PRY kl-3 kl-2 Pp1-Y1 ARY Ppr-Y WDY Pp1-Y2 ORY CCY Dmau kl-5 Pp1-Y1 ORY kl-2 CCY Pp1-Y2 PRY kl-3 Ppr-Y ARY WDY Dsim kl-5 Pp1-Y1 Pp1-Y2 ORY kl-3 PRY Ppr-Y ARY kl-2 CCY WDY Dsec kl-5 Pp1-Y1 Pp1-Y2 PRY ORY kl-2 Ppr-Y ARY CCY kl-3 WDY

23 Gene duplications Most genes have duplicate copies Degenerated copies 10.0 Dmel Dmau Dsim Dsec ARY copy number complete degenerated exon number

24 Gene duplications Most genes have duplicate copies Degenerated copies 15 Dmel Dmau Dsim Dsec kl-2 copy number 10 5 complete degenerated complete degenerated exon number

25 New genes in simulans clade 2 Mya 0.24 Mya To Y Photo: A. Karwath simulans clade

26 New genes in simulans clade 2 Mya 0.24 Mya CG3511 (2R) Dmel Dsim Dsec Dmau CG3511

27 New genes in simulans clade e.g. Dmau (2R) parent gene CG3511 Dmau Y-linked copies: all inactivated Invader2

28 New genes in simulans clade 2 Mya 0.24 Mya Hrb27C (2L), CG16781 (X), SRPK (2R) CG3511 (2R) Dmel Dsim Dsec Dmau CG3511 Hrb27C CG16781 SRPK inactivated

29 Comparative Y chromosome evolution in Drosophila I. Y chromosome assembly % of Y chromosomes in contigs 2L 2R 3L 3R 4 X Y II. Y-linked gene evolution -Reorganization -Duplication -New genes -Subfunctionalization Dmel Dmau

30 Acknowledgments Ching-Ho Chang (U. Rochester) Casey Bergman (UGA) sim clade PacBio genomes Mahul Chakorbharty (UC Irvine) J.J. Emerson (UC Irvine) Kristi Montooth (U Nebraska) Colin Meiklejohn (U Nebraska) Jeffrey Vedenayagam (NYU) Center for Integrated Research Computing