1 The impact of recombination hotspots on genome evolution of 1 a fungal plant pathogen 2 3 Daniel Croll *,§ , Mark H. Lendenmann * , Ethan Stewart * , Bruce A. McDonald * 4 5 Affiliations 6 * Plant Pathology, Institute of Integrative Biology, ETH Zurich, 8092 Zurich, Switzerland 7 § Michael Smith Laboratories, University of British Columbia, V6T 1Z4 Vancouver BC, 8 Canada 9 10 11 ACCESSION NUMBERS 12 NCBI BioSample accessions for parental strains are SRS383146, SRS383147, SRS383142 and 13 SRS383143. Progeny sequence data is deposited under NCBI BioProject accession numbers 14 PRJNA256988 (cross ST99CH3D1 x ST99CH3D7) and PRJNA256991 (cross ST99CH1A5 x ST99CH1E4). 15 16 17 SHORT TITLE 18 Recombination hotspot variability 19 20 21 Genetics: Early Online, published on September 21, 2015 as 10.1534/genetics.115.180968 Copyright 2015.
48
Embed
The impact of recombination hotspots on genome … · 79 role in disease outbreaks caused by viruses, ... 80 Epidemic influenza is driven by annual re-occurring outbreaks of recombined
This document is posted to help you gain knowledge. Please leave a comment to let me know what you think about it! Share it to your friends and learn new things together.
Transcript
1
The impact of recombination hotspots on genome evolution of 1
a fungal plant pathogen 2
3
Daniel Croll*,§, Mark H. Lendenmann*, Ethan Stewart*, Bruce A. McDonald* 4
5
Affiliations 6
* Plant Pathology, Institute of Integrative Biology, ETH Zurich, 8092 Zurich, Switzerland 7
§ Michael Smith Laboratories, University of British Columbia, V6T 1Z4 Vancouver BC, 8
Canada 9
10
11
ACCESSION NUMBERS 12
NCBI BioSample accessions for parental strains are SRS383146, SRS383147, SRS383142 and 13
SRS383143. Progeny sequence data is deposited under NCBI BioProject accession numbers 14
PRJNA256988 (cross ST99CH3D1 x ST99CH3D7) and PRJNA256991 (cross ST99CH1A5 x ST99CH1E4). 15
16
17
SHORT TITLE 18
Recombination hotspot variability 19
20
21
Genetics: Early Online, published on September 21, 2015 as 10.1534/genetics.115.180968
� 13), quality by depth (QD � 5), read position rank sum test (ReadPosRankSumTest � -8) 538
and Fisher's Exact Test for strand bias (FS � 40). After a joint SNP locus filtering, we 539
filtered genotypes for each parental isolate and progeny at each retained locus. We 540
required that each included isolate had a minimum mean genotyping depth of 5 high-541
24
quality reads. We retained individual genotypes if the phred-scaled genotype quality (GQ) 542
assigned by GATK UnifiedGenotyper was at least 30. 543
544
Genetic map construction and quality assessment 545
We constructed a genotype matrix containing all progeny that were genotyped at a 546
minimum of 90% of all SNPs. Subsequently, we removed all SNP markers that were 547
genotyped in less than 90% of the progeny. We assessed the clonal fraction among the 548
progeny in the offspring populations using the r/qtl package in R (ARENDS et al. 2010). If 549
two progeny were identical at 90% or more of the SNPs, we randomly excluded one of the 550
two likely clones. We inspected the quality of the progeny genotype set for problematic 551
double crossovers at very closely spaced markers. For this, we calculated error LOD 552
scores (LINCOLN and LANDER 1992) as implemented in the r/qtl package. The error LOD 553
score compares likelihoods for a correct genotype versus an erroneous genotype based 554
on pairwise linkage. We excluded genotypes exceeding an error LOD score of 2 from the 555
dataset. We filtered the progeny genotype data for random genotyping errors. We required 556
that any double crossover event in a progeny must be spanned by at least three 557
consecutive SNPs. Furthermore, any double crossover event had to span at least 500 bp. 558
These filtering steps ruled out that double crossover events could be erroneously 559
produced by a single stack (i.e. restriction site) of RAD sequencing reads. In addition, we 560
visually inspected the genotype grid for potential erroneous read mapping or translocation 561
events indicated by switched genotypes in all progeny of a cross. We identified three 562
potential erroneous locations in the cross of 3D1x3D7, with each spanning a maximum 563
physical distance of 650 bp. In the 1A5x1E4 cross, we found three potential erroneous 564
locations with SNP loci spanning a maximum of 50 bp for each region. SNP loci within an 565
erroneous mapping location were excluded from further analyses. 566
567
25
In summary, we retained 23,563 SNPs in 227 progeny of the cross between parental 568
isolates 3D1 and 3D7. We analyzed 214 progeny at 23,284 SNP markers for the cross 569
between parental isolates 1A5 and 1E4. The SNP marker density was 0.6 SNP per kb. 570
The genotyping rate averaged 96.5% (chromosomal averages 93.7 - 97.3%) for cross 571
3D1x3D7 and 95.0% (chromosomal averages 90.9 - 98.3%) for cross 1A5x1E4 (Table 1). 572
SNP markers covered the entire length of all core chromosomes with the exception of 573
telomeric regions. Marker densities were similarly distributed along chromosomes in both 574
crosses. 575
We calculated recombination fractions among all pairs of retained markers using the r/qtl 576
package. We set the maximum of iterations to 10,000 and the tolerance for recombination 577
fraction estimates to 0.0001. Using identical settings, we calculated genetic distances for 578
marker pairs on each chromosome. 579
580
Genetic map reconstruction of chromosome 13 581
Based on pairwise recombination fractions, we identified one problematic region located 582
on chromosome 13 (Figure S1). In both crosses, the pattern of recombination fractions 583
suggested that the parental isolates differed from the reference genome used for read 584
mapping. In order to generate the correct marker order on chromosome 13, we produced 585
the genetic map using r/rqtl. We used 10,000 iterations and a tolerance of 0.0001 for cM 586
estimates between markers. The re-calculated marker order showed that the problematic 587
region contains a large weakly recombining region (Figure S1). As the marker order in 588
weakly recombining regions is difficult to confirm with a high confidence, we excluded 589
chromosome 13 from analyses requiring exact physical locations of markers. 590
591
26
Correlations of recombination rates and genome characteristics 592
We downloaded the reference genome IPO323 (assembly version MG2, Sep 2008; 593
(GOODWIN et al. 2011)) from http://genome.jgi-psf.org/Mycgr3/Mycgr3.home.html 594
(accessed March 2014). Gene annotations of Z. tritici related to the RefSeq assembly ID 595
GCF_000219625.1 were accessed in January 2014. SignalP annotations for gene models 596
of IPO323 were retrieved from http://genome.jgi-psf.org/Mycgr3/Mycgr3.home.html 597
(accessed March 2014). Enrichment for signalP secretion signals (probability > 0.9) was 598
tested using a hypergeometric test in R. 599
600
Motif search in recombination hotspots 601
We searched recombination hotspots for enriched sequence motifs with HOMER v 4.3 602
(HEINZ et al. 2010). For this, we divided the chromosome sequences into non-overlapping 603
10 kb segments. For each cross, we identified segments showing at least 10 crossover 604
events. We used the HOMER findMotifsGenome module to identify enriched oligo-605
nucleotides in the hotspot segments compared to the genomic background divided into 606
equally long segments of 10 kb. We used GC-content auto-normalization to correct for 607
bias in sequence composition. The motif length search was restricted to 8, 10 and 12 bp. 608
The top scoring motif was then used to detect chromosomal regions containing the motif 609
using the annotatePeaks tool included in Homer. 610
611 Population genomic analyses of linkage disequilibrium 612
We analyzed Illumina whole-genome sequence data from all four parental isolates and 21 613
additional isolates (data available under NCBI BioProject PRJNA178194) from the same 614
Swiss population (TORRIANI et al. 2011). We used identical reference genome alignment 615
and SNP quality filtering procedures as described above for the RADseq analyses. In 616
order to correlate recombination rates and linkage disequilibria, we retained SNP positions 617
for the population dataset if these positions were included in the genetic map construction. 618
27
Adjacent SNP marker pairs were omitted if markers were separated by less than 500 bp. 619
All linkage disequilibria calculations were made with vcftools v. 0.1.12a (DANECEK et al. 620
2011). 621
622
623 Acknowledgements 624 625
We are grateful to Christine Grossen, Sam Yeaman, Alan Brelsford and Jessica Purcell for 626
helpful comments on previous versions of the manuscript. The Genetic Diversity Center 627
and the Quantitative Genomics Facility at ETH Zurich were used to generate sequence 628
data. Funding was provided by Swiss National Science Foundation grants to DC 629
[PA00P3_145360], BAM [31003A_134755] and an ETH Zurich grant [ETH-03 12] to DC 630
and BAM.631
28
References 632
633
1000 GENOMES PROJECT CONSORTIUM, ABECASIS G. R., ALTSHULER D., AUTON A., BROOKS L. D., 634 DURBIN R. M., GIBBS R. A., HURLES M. E., MCVEAN G. A., 2010 A map of human genome 635 variation from population-scale sequencing. Nature 467: 1061–1073. 636
AKHUNOV E. D., GOODYEAR A. W., GENG S., QI L.-L., ECHALIER B., GILL B. S., MIFTAHUDIN, 637 GUSTAFSON J. P., LAZO G., CHAO S., ANDERSON O. D., LINKIEWICZ A. M., DUBCOVSKY J., LA 638 ROTA M., SORRELLS M. E., ZHANG D., NGUYEN H. T., KALAVACHARLA V., HOSSAIN K., KIANIAN S. 639 F., PENG J., LAPITAN N. L. V., GONZALEZ-HERNANDEZ J. L., ANDERSON J. A., CHOI D.-W., CLOSE 640 T. J., DILBIRLIGI M., GILL K. S., WALKER-SIMMONS M. K., STEBER C., MCGUIRE P. E., QUALSET 641 C. O., DVORAK J., 2003 The organization and rate of evolution of wheat genomes are 642 correlated with recombination rates along chromosome arms. Genome Res 13: 753–763. 643
ANDERSON L. K., DOYLE G. G., BRIGHAM B., CARTER J., HOOKER K. D., LAI A., RICE M., STACK S. 644 M., 2003 High-resolution crossover maps for each bivalent of Zea mays using recombination 645 nodules. Genetics 165: 849–865. 646
ARENDS D., PRINS P., JANSEN R. C., BROMAN K. W., 2010 R/qtl: high-throughput multiple QTL 647 mapping. Bioinformatics 26: 2990–2992. 648
ARNHEIM N., CALABRESE P., TIEMANN-BOEGE I., 2007 Mammalian meiotic recombination hot 649 spots. Annual Review of Genetics 41: 369–399. 650
AUTON A., FLEDEL-ALON A., PFEIFER S., VENN O., SÉGUREL L., STREET T., LEFFLER E. M., BOWDEN 651 R., ANEAS I., BROXHOLME J., HUMBURG P., IQBAL Z., LUNTER G., MALLER J., HERNANDEZ R. D., 652 MELTON C., VENKAT A., NOBREGA M. A., BONTROP R., MYERS S., DONNELLY P., PRZEWORSKI 653 M., MCVEAN G., 2012 A fine-scale chimpanzee genetic map from population sequencing. 654 Science 336: 193–198. 655
AWADALLA P., 2003 The evolutionary genomics of pathogen recombination. Nat Rev Genet 4: 50–656 60. 657
BACKSTRÖM N., FORSTMEIER W., SCHIELZETH H., MELLENIUS H., NAM K., BOLUND E., WEBSTER M. 658 T., OST T., SCHNEIDER M., KEMPENAERS B., ELLEGREN H., 2010 The recombination landscape 659 of the zebra finch Taeniopygia guttata genome. Genome Res 20: 485–495. 660
BAIRD N. A., ETTER P. D., ATWOOD T. S., CURREY M. C., SHIVER A. L., LEWIS Z. A., SELKER E. U., 661 CRESKO W. A., JOHNSON E. A., 2008 Rapid SNP discovery and genetic mapping using 662 sequenced RAD markers. PLoS ONE 3: e3376. 663
BAKER B. S., CARPENTER A. T., ESPOSITO M. S., ESPOSITO R. E., SANDLER L., 1976 The genetic 664 control of meiosis. Annual Review of Genetics 10: 53–134. 665
BARRETT L. G., THRALL P. H., DODDS P. N., LINDE C. C., 2009 Diversity and evolution of effector 666 loci in natural populations of the plant pathogen Melampsora lini. Mol Biol Evol 26: 2499–2513. 667
BARTON A. B., PEKOSZ M. R., KURVATHI R. S., KABACK D. B., 2008 Meiotic Recombination at the 668 Ends of Chromosomes in Saccharomyces cerevisiae. Genetics 179: 1221–1235. 669
BEGUN D. J., AQUADRO C. F., 1992 Levels of naturally occurring DNA polymorphism correlate with 670 recombination rates in D. melanogaster. Nature 356: 519–520. 671
BOLGER A. M., LOHSE M., USADEL B., 2014 Trimmomatic: a flexible trimmer for Illumina sequence 672 data. Bioinformatics. 673
29
BRADLEY K. M., BREYER J. P., MELVILLE D. B., BROMAN K. W., KNAPIK E. W., SMITH J. R., 2011 An 674 SNP-Based Linkage Map for Zebrafish Reveals Sex Determination Loci. G3 (Bethesda) 1: 3–675 9. 676
BROWN P. W., JUDIS L., CHAN E. R., SCHWARTZ S., SEFTEL A., THOMAS A., HASSOLD T. J., 2005 677 Meiotic synapsis proceeds from a limited number of subtelomeric sites in the human male. Am 678 J Hum Genet 77: 556–566. 679
BRUNNER P. C., TORRIANI S. F. F., CROLL D., STUKENBROCK E. H., MCDONALD B. A., 2013 680 Coevolution and life cycle specialization of plant cell wall degrading enzymes in a 681 hemibiotrophic pathogen. Mol Biol Evol 30: 1337–1347. 682
BULL J. J., 1983 Evolution of sex determining mechanisms. The Benjamin/Cummings Publishing 683 Company, Inc. 684
BYRNES E. J., LI W., REN P., LEWIT Y., VOELZ K., FRASER J. A., DIETRICH F. S., MAY R. C., 685 CHATURVEDI S., CHATUVERDI S., CHATURVEDI V., CHATUVERDI V., HEITMAN J., 2011 A diverse 686 population of Cryptococcus gattii molecular type VGIII in southern Californian HIV/AIDS 687 patients. PLoS Pathog 7: e1002205. 688
CHARLESWORTH B., CHARLESWORTH D., 2000 The degeneration of Y chromosomes. Philos Trans 689 R Soc Lond, B, Biol Sci 355: 1563–1572. 690
CHERRY J. M., BALL C., WENG S., JUVIK G., SCHMIDT R., ADLER C., DUNN B., DWIGHT S., RILES L., 691 MORTIMER R. K., BOTSTEIN D., 1997 Genetic and physical maps of Saccharomyces 692 cerevisiae. Nature 387: 67–73. 693
COOP G., MYERS S. R., 2007 Live hot, die young: transmission distortion in recombination 694 hotspots. PLoS Genet 3: e35. 695
COWGER C., BRUNNER P. C., MUNDT C. C., 2008 Frequency of sexual recombination by 696 Mycosphaerella graminicola in mild and severe epidemics. Phytopathology 98: 752–759. 697
COWGER C., MCDONALD B. A., MUNDT C. C., 2002 Frequency of Sexual Reproduction by 698 Mycosphaerella graminicola on Partially Resistant Wheat Cultivars. Phytopathology 92: 1175–699 1181. 700
CROLL D., MCDONALD B. A., 2012 The accessory genome as a cradle for adaptive evolution in 701 pathogens. PLoS Pathog 8: e1002608. 702
CROLL D., ZALA M., MCDONALD B. A., 2013 Breakage-fusion-bridge cycles and large insertions 703 contribute to the rapid evolution of accessory chromosomes in a fungal pathogen. PLoS Genet 704 9: e1003567. 705
CUTTER A. D., PAYSEUR B. A., 2013 Genomic signatures of selection at linked sites: unifying the 706 disparity among species. Nat Rev Genet 14: 262–274. 707
DANECEK P., AUTON A., ABECASIS G., ALBERS C. A., BANKS E., DEPRISTO M. A., HANDSAKER R. E., 708 LUNTER G., MARTH G. T., SHERRY S. T., MCVEAN G., DURBIN R., 1000 GENOMES PROJECT 709 ANALYSIS GROUP, 2011 The variant call format and VCFtools. Bioinformatics 27: 2156–2158. 710
DEPRISTO M. A., BANKS E., POPLIN R., GARIMELLA K. V., MAGUIRE J. R., HARTL C., PHILIPPAKIS A. 711 A., DEL ANGEL G., RIVAS M. A., HANNA M., MCKENNA A., FENNELL T. J., KERNYTSKY A. M., 712 SIVACHENKO A. Y., CIBULSKIS K., GABRIEL S. B., ALTSHULER D., DALY M. J., 2011 A framework 713 for variation discovery and genotyping using next-generation DNA sequencing data. Nat Genet 714 43: 491–498. 715
DIDELOT X., ACHTMAN M., PARKHILL J., THOMSON N. R., FALUSH D., 2007 A bimodal pattern of 716
30
relatedness between the Salmonella Paratyphi A and Typhi genomes: Convergence or 717 divergence by homologous recombination? Genome Res 17: 61–68. 718
DROUAUD J., 2005 Variation in crossing-over rates across chromosome 4 of Arabidopsis thaliana 719 reveals the presence of meiotic recombination “hot spots.” Genome Res 16: 106–114. 720
DURET L., ARNDT P. F., 2008 The impact of recombination on nucleotide substitutions in the 721 human genome. PLoS Genet 4: e1000071. 722
DURET L., GALTIER N., 2009 Biased gene conversion and the evolution of mammalian genomic 723 landscapes. Annu Rev Genom Hum G 10: 285–311. 724
ETTER P. D., BASSHAM S., HOHENLOHE P. A., JOHNSON E. A., CRESKO W. A., 2011 SNP discovery 725 and genotyping for evolutionary genetics using RAD sequencing. Methods Mol. Biol. 772: 157–726 178. 727
FARRER R. A., WEINERT L. A., BIELBY J., GARNER T. W. J., BALLOUX F., CLARE F., BOSCH J., 728 CUNNINGHAM A. A., WELDON C., PREEZ DU L. H., ANDERSON L., POND S. L. K., SHAHAR-GOLAN 729 R., HENK D. A., FISHER M. C., 2011 Multiple emergences of genetically diverse amphibian-730 infecting chytrids include a globalized hypervirulent recombinant lineage. Proc Natl Acad Sci 731 USA 108: 18732–18736. 732
FISHER M. C., GARNER T. W. J., WALKER S. F., 2009 Global emergence of Batrachochytrium 733 dendrobatidis and amphibian chytridiomycosis in space, time, and host. Annu Rev Microbiol 734 63: 291–310. 735
FRASER J. A., GILES S. S., WENINK E. C., GEUNES-BOYER S. G., WRIGHT J. R., DIEZMANN S., ALLEN 736 A., STAJICH J. E., DIETRICH F. S., PERFECT J. R., HEITMAN J., 2005 Same-sex mating and the 737 origin of the Vancouver Island Cryptococcus gattii outbreak. Nature 437: 1360–1364. 738
GERTON J. L., DERISI J., SHROFF R., LICHTEN M., BROWN P. O., PETES T. D., 2000 Global mapping 739 of meiotic recombination hotspots and coldspots in the yeast Saccharomyces cerevisiae. Proc 740 Natl Acad Sci USA 97: 11383–11390. 741
GOODWIN S. B., M'BAREK S. B., WITTENBERG A. H. J., CRANE C. F., HANE J. K., VAN DER LEE T. A. 742 J., GRIMWOOD J., AERTS A., ANTONIW J., BOWLER J., VAN DER BURGT A., COUTINHO P. M., 743 CSUKAI M., DEHAL P., HAMMOND-KOSACK K. E., HENRISSAT B., KILIAN A., LINDQUIST E., 744 MEHRABI R., RUDD J. J., SALAMOV A., SCHMUTZ J., SCHOUTEN H. J., SHAPIRO H., 745 STERGIOPOULOS I., TORRIANI S. F. F., DE VRIES R. P., WAALWIJK C., WARE S. B., ZWIERS L.-H., 746 OLIVER R. P., GRIGORIEV I. V., KEMA G. H. J., 2011 Finished genome of the fungal wheat 747 pathogen Mycosphaerella graminicola reveals dispensome structure, chromosome plasticity, 748 and stealth pathogenesis. PLoS Genet 7: e1002070. 749
GRIGG M. E., BONNEFOY S., HEHL A. B., SUZUKI Y., BOOTHROYD J. C., 2001 Success and 750 virulence in Toxoplasma as the result of sexual recombination between two distinct ancestries. 751 Science 294: 161–165. 752
HAMILTON W. D., 1980 Sex versus non-sex versus parasite. Oikos 35: 282–290. 753
HASSOLD T., HUNT P., 2001 To ERR (meiotically) is human: The genesis of human aneuploidy. 754 Nat Rev Genet 2: 280–291. 755
HEINZ S., BENNER C., SPANN N., BERTOLINO E., LIN Y. C., LASLO P., CHENG J. X., MURRE C., SINGH 756 H., GLASS C. K., 2010 Simple combinations of lineage-determining transcription factors prime 757 cis-regulatory elements required for macrophage and B cell identities. Mol. Cell 38: 576–589. 758
HILL W. G., ROBERTSON A., 1966 The effect of linkage on limits to artificial selection. Genetical 759 Research 8: 269–294. 760
31
HOLT K. E., PARKHILL J., MAZZONI C. J., ROUMAGNAC P., WEILL F.-X., GOODHEAD I., RANCE R., 761 BAKER S., MASKELL D. J., WAIN J., DOLECEK C., ACHTMAN M., DOUGAN G., 2008 High-762 throughput sequencing provides insights into genome variation and evolution in Salmonella 763 Typhi. Nat Genet 40: 987–993. 764
HURLES M., 2005 How homologous recombination generates a mutable genome. Hum. Genomics 765 2: 179–186. 766
JENSEN-SEAMAN M. I., FUREY T. S., PAYSEUR B. A., LU Y., ROSKIN K. M., CHEN C.-F., THOMAS M. 767 A., HAUSSLER D., JACOB H. J., 2004 Comparative recombination rates in the rat, mouse, and 768 human genomes. Genome Res 14: 528–538. 769
JIANG H., LI N., GOPALAN V., ZILVERSMIT M. M., VARMA S., NAGARAJAN V., LI J., MU J., HAYTON K., 770 HENSCHEN B., YI M., STEPHENS R., MCVEAN G., AWADALLA P., WELLEMS T. E., SU X.-Z., 2011 771 High recombination rates and hotspots in a Plasmodium falciparum genetic cross. Genome 772 Biol 12: R33. 773
KABACK D. B., BARBER D., MAHON J., LAMB J., YOU J., 1999 Chromosome size-dependent control 774 of meiotic reciprocal recombination in Saccharomyces cerevisiae: the role of crossover 775 interference. Genetics 152: 1475–1486. 776
KARASOV T. L., HORTON M. W., BERGELSON J., 2014 Genomic variability as a driver of plant-777 pathogen coevolution? Curr Opin Plant Biol 18C: 24–30. 778
KEMA G. H. J., GOODWIN S. B., HAMZA S., VERSTAPPEN E. C. P., CAVALETTO J. R., VAN DER LEE T. 779 A. J., DE WEERDT M., BONANTS P. J. M., WAALWIJK C., 2002 A combined amplified fragment 780 length polymorphism and randomly amplified polymorphism DNA genetic kinkage map of 781 Mycosphaerella graminicola, the septoria tritici leaf blotch pathogen of wheat. Genetics 161: 782 1497–1505. 783
KEMA G. H. J., VERSTAPPEN E. C. P., TODOROVA M., WAALWIJK C., 1996 Successful crosses and 784 molecular tetrad and progeny analyses demonstrate heterothallism in Mycosphaerella 785 graminicola. Curr Genet 30: 251–258. 786
KONG A., GUDBJARTSSON D. F., SAINZ J., JONSDOTTIR G. M., GUDJONSSON S. A., RICHARDSSON B., 787 SIGURDARDOTTIR S., BARNARD J., HALLBECK B., MASSON G., SHLIEN A., PALSSON S. T., FRIGGE 788 M. L., THORGEIRSSON T. E., GULCHER J. R., STEFANSSON K., 2002 A high-resolution 789 recombination map of the human genome. Nat Genet. 790
LAMBREGHTS R., SHI M., BELDEN W. J., DECAPRIO D., PARK D., HENN M. R., GALAGAN J. E., 791 BAŞTÜRKMEN M., BIRREN B. W., SACHS M. S., DUNLAP J. C., LOROS J. J., 2009 A high-density 792 single nucleotide polymorphism map for Neurospora crassa. Genetics 181: 767–781. 793
LANGMEAD B., SALZBERG S. L., 2012 Fast gapped-read alignment with Bowtie 2. Nat Methods 9: 794 357–359. 795
LINCOLN S. E., LANDER E. S., 1992 Systematic detection of errors in genetic linkage data. 796 Genomics 14: 604–610. 797
LIVELY C. M., 2010 A Review of Red Queen Models for the Persistence of Obligate Sexual 798 Reproduction. J Hered 101: S13–S20. 799
MANCERA E., BOURGON R., BROZZI A., HUBER W., STEINMETZ L. M., 2008 High-resolution mapping 800 of meiotic crossovers and non-crossovers in yeast. Nature 454: 479–485. 801
MCDONALD B. A., LINDE C., 2002 Pathogen population genetics, evolutionary potential, and 803
32
durable resistance. Annu Rev Phytopathol 40: 349–379. 804
MENKIS A., JACOBSON D. J., GUSTAFSSON T., JOHANNESSON H., 2008 The mating-type 805 chromosome in the filamentous ascomycete Neurospora tetrasperma represents a model for 806 early evolution of sex chromosomes. PLoS Genet 4: e1000030. 807
MIRZADI GOHARI A., MEHRABI R., ROBERT O., INCE I. A., BOEREN S., SCHUSTER M., STEINBERG G., 808 DE WIT P. J. G. M., KEMA G. H. J., 2014 Molecular characterization and functional analyses of 809 ZtWor1, a transcriptional regulator of the fungal wheat pathogen Zymoseptoria tritici. Mol Plant 810 Pathol 15: 394–405. 811
MORAIS DO AMARAL A., ANTONIW J., RUDD J. J., HAMMOND-KOSACK K. E., 2012 Defining the 812 Predicted Protein Secretome of the Fungal Wheat Leaf Pathogen Mycosphaerella graminicola. 813 PLoS ONE 7: e49904. 814
MORRAN L. T., SCHMIDT O. G., GELARDEN I. A., PARRISH R. C., LIVELY C. M., 2011 Running with 815 the Red Queen: host-parasite coevolution selects for biparental sex. Science 333: 216–218. 816
MORTIMER R. K., SCHILD D., CONTOPOULOU C. R., KANS J. A., 1989 Genetic map of 817 Saccharomyces cerevisiae, edition 10. Yeast 5: 321–403. 818
MUYLE A., SERRES-GIARDI L., RESSAYRE A., ESCOBAR J., GLÉMIN S., 2011 GC-biased gene 819 conversion and selection affect GC content in the Oryza genus (rice). Mol Biol Evol 28: 2695–820 2706. 821
MYERS S., BOTTOLO L., FREEMAN C., MCVEAN G., DONNELLY P., 2005 A fine-scale map of 822 recombination rates and hotspots across the human genome. Science 310: 321–324. 823
MYERS S., FREEMAN C., AUTON A., DONNELLY P., MCVEAN G., 2008 A common sequence motif 824 associated with recombination hot spots and genome instability in humans. Nat Genet 40: 825 1124–1129. 826
NACHMAN M. W., 2002 Variation in recombination rate across the genome: evidence and 827 implications. Curr. Opin. Genet. Dev. 12: 657–663. 828
NARANJO T., CORREDOR E., 2008 Nuclear architecture and chromosome dynamics in the search 829 of the pairing partner in meiosis in plants. Cytogenet Genome Res 120: 320–330. 830
NELSON M. I., HOLMES E. C., 2007 The evolution of epidemic influenza. Nat Rev Genet 8: 196–831 205. 832
O'DRISCOLL A., KILDEA S., DOOHAN F., SPINK J., 2014 The wheat–Septoria conflict: a new front 833 opening up? Trends in plant …. 834
OTTO S. P., BARTON N. H., 1997 The evolution of recombination: removing the limits to natural 835 selection. Genetics 147: 879–906. 836
OTTO S. P., LENORMAND T., 2002 Resolving the paradox of sex and recombination. Nat Rev 837 Genet 3: 252–261. 838
PETRE B., KAMOUN S., 2014 How do filamentous pathogens deliver effector proteins into plant 839 cells? PLoS Biol 12: e1001801. 840
RAFFAELE S., KAMOUN S., 2012 Genome evolution in filamentous plant pathogens: why bigger 841 can be better. Nat Rev Microbiol 10: 417–430. 842
RAFFAELE S., FARRER R. A., CANO L. M., STUDHOLME D. J., MACLEAN D., THINES M., JIANG R. H. Y., 843 ZODY M. C., KUNJETI S. G., DONOFRIO N. M., MEYERS B. C., NUSBAUM C., KAMOUN S., 2010 844
33
Genome evolution following host jumps in the Irish potato famine pathogen lineage. Science 845 330: 1540–1543. 846
ROESTI M., MOSER D., BERNER D., 2013 Recombination in the threespine stickleback genome--847 patterns and consequences. Molecular Ecology 22: 3014–3027. 848
ROSENBLUM E. B., STAJICH J. E., MADDOX N., EISEN M. B., 2008 Global gene expression profiles 849 for life stages of the deadly amphibian pathogen Batrachochytrium dendrobatidis. Proc Natl 850 Acad Sci USA 105: 17034–17039. 851
STAJICH J. E., WILKE S. K., AHRÉN D., AU C. H., BIRREN B. W., BORODOVSKY M., BURNS C., 852 CANBÄCK B., CASSELTON L. A., CHENG C. K., DENG J., DIETRICH F. S., FARGO D. C., FARMAN M. 853 L., GATHMAN A. C., GOLDBERG J., GUIGÓ R., HOEGGER P. J., HOOKER J. B., HUGGINS A., JAMES 854 T. Y., KAMADA T., KILARU S., KODIRA C., KÜES U., KUPFER D., KWAN H. S., LOMSADZE A., LI W., 855 LILLY W. W., MA L.-J., MACKEY A. J., MANNING G., MARTIN F., MURAGUCHI H., NATVIG D. O., 856 PALMERINI H., RAMESH M. A., REHMEYER C. J., ROE B. A., SHENOY N., STANKE M., TER-857 HOVHANNISYAN V., TUNLID A., VELAGAPUDI R., VISION T. J., ZENG Q., ZOLAN M. E., PUKKILA P. 858 J., 2010 Insights into evolution of multicellular fungi from the assembled chromosomes of the 859 mushroom Coprinopsis cinerea (Coprinus cinereus). Proc Natl Acad Sci USA 107: 11889–860 11894. 861
STEINER W. W., SMITH G. R., 2005 Natural meiotic recombination hot spots in the 862 Schizosaccharomyces pombe genome successfully predicted from the simple sequence motif 863 M26. Mol Cell Biol 25: 9054–9062. 864
STUKENBROCK E. H., MCDONALD B. A., 2008 The origins of plant pathogens in agro-ecosystems. 865 Annu Rev Phytopathol 46: 75–100. 866
THRALL P. H., LAINE A.-L., RAVENSDALE M., NEMRI A., DODDS P. N., BARRETT L. G., BURDON J. J., 867 2012 Rapid genetic change underpins antagonistic coevolution in a natural host-pathogen 868 metapopulation. Ecol Lett 15: 425–435. 869
TORRIANI S. F. F., STUKENBROCK E. H., BRUNNER P. C., MCDONALD B. A., CROLL D., 2011 870 Evidence for extensive recent intron transposition in closely related fungi. Curr Biol 21: 2017–871 2022. 872
TORRIANI S. F., BRUNNER P. C., MCDONALD B. A., SIEROTZKI H., 2009 QoI resistance emerged 873 independently at least 4 times in European populations of Mycosphaerella graminicola. Pest 874 Manag. Sci. 65: 155–162. 875
TSAI I. J., BURT A., KOUFOPANOU V., 2010 Conservation of recombination hotspots in yeast. Proc 876 Natl Acad Sci USA 107: 7847–7852. 877
WEBSTER M. T., HURST L. D., 2012 Direct and indirect consequences of meiotic recombination: 878 implications for genome evolution. Trends Genet 28: 101–109. 879
WENDTE J. M., MILLER M. A., LAMBOURN D. M., MAGARGAL S. L., JESSUP D. A., GRIGG M. E., 2010 880 Self-mating in the definitive host potentiates clonal outbreaks of the apicomplexan parasites 881 Sarcocystis neurona and Toxoplasma gondii. PLoS Genet 6: e1001261. 882
WILSON M. A., MAKOVA K. D., 2009 Genomic analyses of sex chromosome evolution. Annu Rev 883 Genom Hum G 10: 333–354. 884
WITTENBERG A. H. J., VAN DER LEE T. A. J., BEN M'BAREK S., WARE S. B., GOODWIN S. B., KILIAN A., 885 VISSER R. G. F., KEMA G. H. J., SCHOUTEN H. J., 2009 Meiosis drives extraordinary genome 886 plasticity in the haploid fungal plant pathogen Mycosphaerella graminicola. PLoS ONE 4: 887 e5863. 888
34
WU T. C., LICHTEN M., 1995 Factors that affect the location and frequency of meiosis-induced 889 double-strand breaks in Saccharomyces cerevisiae. Genetics 140: 55–66. 890
ZHAN J., KEMA G., WAALWIJK C., MCDONALD B. A., 2002 Distribution of mating type alleles in the 891 wheat pathogen Mycosphaerella graminicola over spatial scales from lesions to continents. 892 Fungal Genet Biol 36: 128–136. 893
ZHAN J., LINDE C. C., JÜRGENS T., MERZ U., STEINEBRUNNER F., MCDONALD B. A., 2005 Variation 894 for neutral markers is correlated with variation for quantitative traits in the plant pathogenic 895 fungus Mycosphaerella graminicola. Molecular Ecology 14: 2683–2693. 896
ZHAN J., MUNDT C. C., MCDONALD B. A., 2007 Sexual reproduction facilitates the adaptation of 897 parasites to antagonistic host environments: Evidence from empirical study in the wheat-898 Mycosphaerella graminicola system. International Journal for Parasitology 37: 861–870. 899
900
901
902
903
35
Figure legends 904 905
Figure 1: Physical distance between all detected recombination events per progeny 906
summarized for each of the two analyzed crosses (3D1x3D7 and 1A5x1E4). A short 907
distance between recombination events in a progeny may indicate a potential 908
noncrossover. In this case the two recombination events would be boundaries of a gene 909
conversion tract. In order to conservatively identify true crossovers, a minimum distance of 910
50 kb was required for a recombination event to be considered a crossover (shaded in 911
black) instead of a putative noncrossover event (shaded in grey). All analyses of 912
recombination rates and hotspots are based on recombination events at least 50 kb from 913
the nearest neighboring recombination event. 914
915
Figure 2: Summary statistics of the genetic maps constructed for the two crosses 916
3D1x3D7 and 1A5x1E4. A) Genetic map length per chromosome. Horizontal hatches 917
show the positions of genetic markers in the genetic maps. In cross 3D1x3D7, four 918
accessory chromosomes were shared between the parental isolates and could therefore 919
be used for map construction. In cross 1A5x1E4, seven accessory chromosomes were 920
shared between the parental isolates. B) Recombination rates per chromosome expressed 921
as cM per Mb. Genetic maps for the two crosses are distinguished by color. See panel C 922
for color legend. C) Number of genetic map positions per chromosome (minimal set of 923
markers separated by at least one crossover). 924
925
Figure 3: Recombination landscape of the plant pathogenic fungus Zymoseptoria 926
tritici. Recombination rates were estimated in two crosses by calculating genetic map 927
distances (in cM) for non-overlapping segments of 20 kb along chromosomes. Variations 928
in recombination rates are shown for cross 1A5x1E4 (blue) and 3D1x3D7 (red). 929
Recombination rates on accessory chromosomes are shown only for chromosomes that 930
36
were found in both parental isolates of a cross. Chromosome 13 contains a region for 931
which recombination rate variations could not be accurately determined (see Methods). 932
933
Figure 4: Recombination rate variations in relation to telomere distance. 934
Recombination rates were calculated in non-overlapping 10 kb segments for core 935
chromosomes. Recombination rates were then summarized into 5% bins representing the 936
relative distance to the telomere end. The boxplot shows the median recombination rate 937
(horizontal bar), the 25% and 75% quartiles as a solid box and the 5% and 95% quantiles 938
as vertical lines. Recombination rates were highest in both crosses for subtelomeric 939
distances of 10-15% of the chromosome length. Core chromosomes vary from 1.19 – 6.09 940
Mb in length. 941
942
Figure 5: Differences in gene density and sequence diversity in the recombination 943
hotspots and non-hotspots of crosses 3D1x3D7 and 1A5x1E4. For both panels the 944
boxplots show the median of the distribution (horizontal bar), the 25% and 75% quartiles 945
as a solid box and the 5% and 95% quantiles as vertical lines. A) Differences in coding 946
sequence density between hotspot segments (each 10 kb) and segments of the genomic 947
background (“non-hotspots”) divided into 10 kb segments. B) Differences in diversity 948
between the two parental genomes (% SNP) assessed for hotspot segments (each 10 kb) 949
and segments of the genomic background (“non-hotspots”) divided into 10 kb segments. 950
951
Figure 6: The top enriched oligonucleotide motifs that were identified by a de novo 952
motif search in the recombination hotspots of crosses 3D1x3D7 and 1A5x1E4. The 953
enrichment of sequence motifs in recombination hotspots was tested using a 954
hypergeometric test. The genomic background was segmented and binned according to 955
GC-content. Enrichment tests were performed using weighted background sequences 956
37
reflecting the GC-content distribution found in hotspots. Weighting background sequences 957
avoided spurious enrichments due to differences in GC-content between the genomic 958
background and hotspots. 959
960
Figure 7: Recombination rates estimated from linkage analyses in crosses 961
compared to linkage disequilibrium estimated from population resequencing data. 962
Recombination rates between adjacent SNP markers were calculated as cM per Mb in 963
crosses 3D1x3D7 and 1A5x1E4. Resequencing data of field isolates from the same 964
regional population as the parents was used to calculate linkage disequilbria (r2). For this, 965
whole genome sequencing data was analyzed for SNPs segregating at identical positions 966
in the genome as in the SNP dataset generated for the two crosses. Different symbols are 967
used to show data points from core and accessory chromosomes. 968
969
38
Tables 970 971
Table 1: Summary of restriction-associated DNA sequencing (RADseq) SNP markers 972
used for the construction of genetic maps. The average genotyping rate of the SNP 973
markers is shown as an average per chromosome and cross. Chromosomes 1-13 are core 974
chromosomes found in all strains of the species, chromosomes 14-21 are accessory. 975
976
977
Cross 3D1x3D7 (227 progeny)
Cross 1A5x1E4 (214 progeny)
Chromosome Number of SNPs Genotyping rate (%) Number of SNPs Genotyping rate (%)
1 4697 96.16 4629 96.15
2 2486 97.26 2290 96.28
3 2299 97.31 2233 96.02
4 1614 96.87 1447 96.08
5 1634 95.62 1778 96.16
6 1398 97.03 1349 96.28
7 1379 97.27 1401 96.16
8 1576 96.54 1537 96.10
9 1448 97.08 1309 95.62
10 1013 97.09 1115 95.95
11 920 97.26 982 96.08
12 923 96.90 775 95.50
13 806 96.95 721 95.94
14 - - 118 90.97
15 - - 461 93.93
16 193 95.49 109 92.37
17 395 93.72 - -
18 - - 110 90.93
19 461 97.06 411 94.13
20 321 94.25 271 94.63
21 - - 238 93.58
978
979
980
39
Table 2: Overview of genetic maps constructed in two crosses of Zymoseptoria 981
tritici. For each chromosome the physical length (in kb), genetic map length (cM) and the 982
number of genetic map positions (minimal number of markers separated by at least one 983
crossover) is shown for each chromosomal map. The recombination rate per chromosome 984