Poly(ADP-ribosyl)ation is a reversible post-translational modification synthetized by ADP-ribose transferases and removed by poly(ADP-ribose) glycohydrolase (PARG), which plays important roles in DNA damage repair. While well-studied in somatic tissues, much less is known about poly(ADP-ribosyl)ation in the germline, where DNA double-strand breaks are introduced by a regulated program and repaired by crossover recombination to establish a tether between homologous chromosomes. The interaction between the parental chromosomes is facilitated by meiotic specific adaptation of the chromosome axes and cohesins, and reinforced by the synaptonemal complex. Here, we uncover an unexpected role for PARG in coordinating the induction of meiotic DNA breaks and their homologous recombination-mediated repair in Caenorhabditis elegans. PARG-1/PARG interacts with both axial and central elements of the synaptonemal complex, REC-8/Rec8 and the MRN/X complex. PARG-1 shapes the recombination landscape and reinforces the tightly regulated control of crossover numbers without requiring its catalytic activity. We unravel roles in regulating meiosis, beyond its enzymatic activity in poly(ADP-ribose) catabolism.

AHN Center for Reproductive Medicine AHN McCandless Pittsburgh PA USA

Bioinformatics and Computational Biology Faculty of Computer Science University of Vienna Vienna Austria

Center for Integrative Bioinformatics Vienna Max Perutz Laboratories Medical University of Vienna Vienna BioCenter University of Vienna Vienna Austria

Centre for Anatomy and Cell Biology Medical University of Vienna Vienna Austria

Department of Biology Faculty of Medicine Masaryk University Brno Czech Republic

Department of Chromosome Biology Max Perutz Laboratories Vienna Biocenter University of Vienna Vienna Austria

Department of Obstetrics Gynecology and Reproductive Sciences Magee Womens Research Institute University of Pittsburgh School of Medicine Pittsburgh PA USA

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Koh DW, et al. Failure to degrade poly(ADP-ribose) causes increased sensitivity to cytotoxicity and early embryonic lethality. Proc. Natl Acad. Sci. U.S.A. 2004;101:17699–17704. doi: 10.1073/pnas.0406182101. PubMed DOI PMC

Menissier de Murcia J, et al. Functional interaction between PARP-1 and PARP-2 in chromosome stability and embryonic development in mouse. EMBO J. 2003;22:2255–2263. doi: 10.1093/emboj/cdg206. PubMed DOI PMC

Slade, D. Mitotic functions of poly(ADP-ribose) polymerases. Biochem. Pharmacol.10.1016/j.bcp.2019.03.028 (2019). PubMed PMC

O’Sullivan J, et al. Emerging roles of eraser enzymes in the dynamic control of protein ADP-ribosylation. Nat. Commun. 2019;10:1182. doi: 10.1038/s41467-019-08859-x. PubMed DOI PMC

Dantzer F, et al. Poly(ADP-ribose) polymerase-2 contributes to the fidelity of male meiosis I and spermiogenesis. Proc. Natl Acad. Sci. U.S.A. 2006;103:14854–14859. doi: 10.1073/pnas.0604252103. PubMed DOI PMC

Ame JC, et al. Radiation-induced mitotic catastrophe in PARG-deficient cells. J. Cell Sci. 2009;122:1990–2002. doi: 10.1242/jcs.039115. PubMed DOI

St-Laurent JF, Gagnon SN, Dequen F, Hardy I, Desnoyers S. Altered DNA damage response in Caenorhabditis elegans with impaired poly(ADP-ribose) glycohydrolases genes expression. DNA Repair (Amst.) 2007;6:329–343. doi: 10.1016/j.dnarep.2006.10.027. PubMed DOI

Byrne, A. B. et al. Inhibiting poly(ADP-ribosylation) improves axon regeneration. Elife5, 10.7554/eLife.12734 (2016). PubMed PMC

Dequen F, Gagnon SN, Desnoyers S. Ionizing radiations in Caenorhabditis elegans induce poly(ADP-ribosyl)ation, a conserved DNA-damage response essential for survival. DNA Repair (Amst.) 2005;4:814–825. doi: 10.1016/j.dnarep.2005.04.015. PubMed DOI

Gagnon SN, Hengartner MO, Desnoyers S. The genes pme-1 and pme-2 encode two poly(ADP-ribose) polymerases in Caenorhabditis elegans. Biochem. J. 2002;368:263–271. doi: 10.1042/bj20020669. PubMed DOI PMC

Zickler D, Kleckner N. Meiotic chromosomes: integrating structure and function. Annu. Rev. Genet. 1999;33:603–754. doi: 10.1146/annurev.genet.33.1.603. PubMed DOI

Zickler, D. & Kleckner, N. Recombination, pairing, and synapsis of homologs during meiosis. Cold Spring Harb. Perspect. Biol.7, 10.1101/cshperspect.a016626 (2015). PubMed PMC

Cao L, Alani E, Kleckner N. A pathway for generation and processing of double-strand breaks during meiotic recombination in S. cerevisiae. Cell. 1990;61:1089–1101. doi: 10.1016/0092-8674(90)90072-M. PubMed DOI

Sun H, Treco D, Schultes NP, Szostak JW. Double-strand breaks at an initiation site for meiotic gene conversion. Nature. 1989;338:87–90. doi: 10.1038/338087a0. PubMed DOI

Keeney S, Giroux CN, Kleckner N. Meiosis-specific DNA double-strand breaks are catalyzed by Spo11, a member of a widely conserved protein family. Cell. 1997;88:375–384. doi: 10.1016/S0092-8674(00)81876-0. PubMed DOI

Chin GM, Villeneuve AM. C. elegans mre-11 is required for meiotic recombination and DNA repair but is dispensable for the meiotic G(2) DNA damage checkpoint. Genes Dev. 2001;15:522–534. doi: 10.1101/gad.864101. PubMed DOI PMC

Meneely PM, McGovern OL, Heinis FI, Yanowitz JL. Crossover distribution and frequency are regulated by him-5 in Caenorhabditis elegans. Genetics. 2012;190:1251–1266. doi: 10.1534/genetics.111.137463. PubMed DOI PMC

Reddy KC, Villeneuve AMC. elegans HIM-17 links chromatin modification and competence for initiation of meiotic recombination. Cell. 2004;118:439–452. doi: 10.1016/j.cell.2004.07.026. PubMed DOI

Wagner CR, Kuervers L, Baillie DL, Yanowitz JL. xnd-1 regulates the global recombination landscape in Caenorhabditis elegans. Nature. 2010;467:839–843. doi: 10.1038/nature09429. PubMed DOI PMC

Stamper EL, et al. Identification of DSB-1, a protein required for initiation of meiotic recombination in Caenorhabditis elegans, illuminates a crossover assurance checkpoint. PLoS Genet. 2013;9:e1003679. doi: 10.1371/journal.pgen.1003679. PubMed DOI PMC

Rosu S, et al. The C. elegans DSB-2 protein reveals a regulatory network that controls competence for meiotic DSB formation and promotes crossover assurance. PLoS Genet. 2013;9:e1003674. doi: 10.1371/journal.pgen.1003674. PubMed DOI PMC

Yin Y, Smolikove S. Impaired resection of meiotic double-strand breaks channels repair to nonhomologous end joining in Caenorhabditis elegans. Mol. Cell. Biol. 2013;33:2732–2747. doi: 10.1128/MCB.00055-13. PubMed DOI PMC

Macaisne N, Kessler Z, Yanowitz JL. Meiotic double-strand break proteins influence repair pathway utilization. Genetics. 2018;210:843–856. doi: 10.1534/genetics.118.301402. PubMed DOI PMC

Serrentino ME, Borde V. The spatial regulation of meiotic recombination hotspots: are all DSB hotspots crossover hotspots? Exp. Cell Res. 2012;318:1347–1352. doi: 10.1016/j.yexcr.2012.03.025. PubMed DOI

Yokoo R, et al. COSA-1 reveals robust homeostasis and separable licensing and reinforcement steps governing meiotic crossovers. Cell. 2012;149:75–87. doi: 10.1016/j.cell.2012.01.052. PubMed DOI PMC

Rosu S, Libuda DE, Villeneuve AM. Robust crossover assurance and regulated interhomolog access maintain meiotic crossover number. Science. 2011;334:1286–1289. doi: 10.1126/science.1212424. PubMed DOI PMC

Youds JL, et al. RTEL-1 enforces meiotic crossover interference and homeostasis. Science. 2010;327:1254–1258. doi: 10.1126/science.1183112. PubMed DOI PMC

Hillers KJ, Villeneuve AM. Chromosome-wide control of meiotic crossing over in C. elegans. Curr. Biol. 2003;13:1641–1647. doi: 10.1016/j.cub.2003.08.026. PubMed DOI

Barnes TM, Kohara Y, Coulson A, Hekimi S. Meiotic recombination, noncoding DNA and genomic organization in Caenorhabditis elegans. Genetics. 1995;141:159–179. PubMed PMC

de los Santos T, et al. The Mus81/Mms4 endonuclease acts independently of double-Holliday junction resolution to promote a distinct subset of crossovers during meiosis in budding yeast. Genetics. 2003;164:81–94. PubMed PMC

Zalevsky J, MacQueen AJ, Duffy JB, Kemphues KJ, Villeneuve AM. Crossing over during Caenorhabditis elegans meiosis requires a conserved MutS-based pathway that is partially dispensable in budding yeast. Genetics. 1999;153:1271–1283. PubMed PMC

Kelly KO, Dernburg AF, Stanfield GM, Villeneuve AM. Caenorhabditis elegans msh-5 is required for both normal and radiation-induced meiotic crossing over but not for completion of meiosis. Genetics. 2000;156:617–630. PubMed PMC

Tsai CJ, et al. Meiotic crossover number and distribution are regulated by a dosage compensation protein that resembles a condensin subunit. Genes Dev. 2008;22:194–211. doi: 10.1101/gad.1618508. PubMed DOI PMC

Colaiacovo MP, et al. Synaptonemal complex assembly in C. elegans is dispensable for loading strand-exchange proteins but critical for proper completion of recombination. Dev. Cell. 2003;5:463–474. doi: 10.1016/S1534-5807(03)00232-6. PubMed DOI

Libuda DE, Uzawa S, Meyer BJ, Villeneuve AM. Meiotic chromosome structures constrain and respond to designation of crossover sites. Nature. 2013;502:703–706. doi: 10.1038/nature12577. PubMed DOI PMC

Pattabiraman D, Roelens B, Woglar A, Villeneuve AM. Meiotic recombination modulates the structure and dynamics of the synaptonemal complex during C. elegans meiosis. PLoS Genet. 2017;13:e1006670. doi: 10.1371/journal.pgen.1006670. PubMed DOI PMC

Machovina TS, et al. A surveillance system ensures crossover formation in C. elegans. Curr. Biol. 2016;26:2873–2884. doi: 10.1016/j.cub.2016.09.007. PubMed DOI PMC

Goodyer W, et al. HTP-3 links DSB formation with homolog pairing and crossing over during C. elegans meiosis. Dev. Cell. 2008;14:263–274. doi: 10.1016/j.devcel.2007.11.016. PubMed DOI

Martinez-Perez E, Villeneuve AM. HTP-1-dependent constraints coordinate homolog pairing and synapsis and promote chiasma formation during C. elegans meiosis. Genes Dev. 2005;19:2727–2743. doi: 10.1101/gad.1338505. PubMed DOI PMC

Zetka MC, Kawasaki I, Strome S, Muller F. Synapsis and chiasma formation in Caenorhabditis elegans require HIM-3, a meiotic chromosome core component that functions in chromosome segregation. Genes Dev. 1999;13:2258–2270. doi: 10.1101/gad.13.17.2258. PubMed DOI PMC

Bae, W., Park, J. H., Lee, M. H., Park, H. W. & Koo, H. S. Hypersensitivity to DNA double-strand breaks associated with PARG deficiency is suppressed by exo-1 and polq-1 mutations in Caenorhabditis elegans. FEBS J. 10.1111/febs.15082 (2019). PubMed

Hodgkin J, Horvitz HR, Brenner S. Nondisjunction mutants of the nematode CAENORHABDITIS ELEGANS. Genetics. 1979;91:67–94. PubMed PMC

Kaufmann T, et al. A novel non-canonical PIP-box mediates PARG interaction with PCNA. Nucleic Acids Res. 2017;45:9741–9759. doi: 10.1093/nar/gkx604. PubMed DOI PMC

Mortusewicz O, Fouquerel E, Ame JC, Leonhardt H, Schreiber V. PARG is recruited to DNA damage sites through poly(ADP-ribose)- and PCNA-dependent mechanisms. Nucleic Acids Res. 2011;39:5045–5056. doi: 10.1093/nar/gkr099. PubMed DOI PMC

Ohashi S, et al. Subcellular localization of poly(ADP-ribose) glycohydrolase in mammalian cells. Biochem. Biophys. Res. Commun. 2003;307:915–921. doi: 10.1016/S0006-291X(03)01272-5. PubMed DOI

Winstall E, et al. Preferential perinuclear localization of poly(ADP-ribose) glycohydrolase. Exp. Cell Res. 1999;251:372–378. doi: 10.1006/excr.1999.4594. PubMed DOI

Meyer-Ficca ML, Meyer RG, Coyle DL, Jacobson EL, Jacobson MK. Human poly(ADP-ribose) glycohydrolase is expressed in alternative splice variants yielding isoforms that localize to different cell compartments. Exp. Cell Res. 2004;297:521–532. doi: 10.1016/j.yexcr.2004.03.050. PubMed DOI

MacQueen AJ, Colaiacovo MP, McDonald K, Villeneuve AM. Synapsis-dependent and -independent mechanisms stabilize homolog pairing during meiotic prophase in C. elegans. Genes Dev. 2002;16:2428–2442. doi: 10.1101/gad.1011602. PubMed DOI PMC

de Carvalho CE, et al. LAB-1 antagonizes the Aurora B kinase in C. elegans. Genes Dev. 2008;22:2869–2885. doi: 10.1101/gad.1691208. PubMed DOI PMC

Martinez-Perez E, et al. Crossovers trigger a remodeling of meiotic chromosome axis composition that is linked to two-step loss of sister chromatid cohesion. Genes Dev. 2008;22:2886–2901. doi: 10.1101/gad.1694108. PubMed DOI PMC

Bhalla N, Wynne DJ, Jantsch V, Dernburg AF. ZHP-3 acts at crossovers to couple meiotic recombination with synaptonemal complex disassembly and bivalent formation in C. elegans. PLoS Genet. 2008;4:e1000235. doi: 10.1371/journal.pgen.1000235. PubMed DOI PMC

Janisiw E, Dello Stritto MR, Jantsch V, Silva N. BRCA1-BARD1 associate with the synaptonemal complex and pro-crossover factors and influence RAD-51 dynamics during Caenorhabditis elegans meiosis. PLoS Genet. 2018;14:e1007653. doi: 10.1371/journal.pgen.1007653. PubMed DOI PMC

Li Q, et al. The tumor suppressor BRCA1-BARD1 complex localizes to the synaptonemal complex and regulates recombination under meiotic dysfunction in Caenorhabditis elegans. PLoS Genet. 2018;14:e1007701. doi: 10.1371/journal.pgen.1007701. PubMed DOI PMC

Jantsch V, et al. Targeted gene knockout reveals a role in meiotic recombination for ZHP-3, a Zip3-related protein in Caenorhabditis elegans. Mol. Cell. Biol. 2004;24:7998–8006. doi: 10.1128/MCB.24.18.7998-8006.2004. PubMed DOI PMC

Paix A, Folkmann A, Rasoloson D, Seydoux G. High efficiency, homology-directed genome editing in caenorhabditis elegans using CRISPR-Cas9 ribonucleoprotein complexes. Genetics. 2015;201:47–54. doi: 10.1534/genetics.115.179382. PubMed DOI PMC

Silva N, et al. The fidelity of synaptonemal complex assembly is regulated by a signaling mechanism that controls early meiotic progression. Dev. Cell. 2014;31:503–511. doi: 10.1016/j.devcel.2014.10.001. PubMed DOI

Crawley O, et al. Cohesin-interacting protein WAPL-1 regulates meiotic chromosome structure and cohesion by antagonizing specific cohesin complexes. Elife. 2016;5:e10851. doi: 10.7554/eLife.10851. PubMed DOI PMC

Rog, O. & Dernburg, A. F. Direct visualization reveals kinetics of meiotic chromosome synapsis. Cell Rep.10.1016/j.celrep.2015.02.032 (2015). PubMed PMC

Alpi A, Pasierbek P, Gartner A, Loidl J. Genetic and cytological characterization of the recombination protein RAD-51 in Caenorhabditis elegans. Chromosoma. 2003;112:6–16. doi: 10.1007/s00412-003-0237-5. PubMed DOI

Lemmens BB, Johnson NM, Tijsterman M. COM-1 promotes homologous recombination during Caenorhabditis elegans meiosis by antagonizing Ku-mediated non-homologous end joining. PLoS Genet. 2013;9:e1003276. doi: 10.1371/journal.pgen.1003276. PubMed DOI PMC

Martin JS, Winkelmann N, Petalcorin MI, McIlwraith MJ, Boulton SJ. RAD-51-dependent and -independent roles of a Caenorhabditis elegans BRCA2-related protein during DNA double-strand break repair. Mol. Cell. Biol. 2005;25:3127–3139. doi: 10.1128/MCB.25.8.3127-3139.2005. PubMed DOI PMC

Petalcorin MI, Galkin VE, Yu X, Egelman EH, Boulton SJ. Stabilization of RAD-51-DNA filaments via an interaction domain in Caenorhabditis elegans BRCA2. Proc. Natl Acad. Sci. U.S.A. 2007;104:8299–8304. doi: 10.1073/pnas.0702805104. PubMed DOI PMC

Penkner A, et al. A conserved function for a Caenorhabditis elegans Com1/Sae2/CtIP protein homolog in meiotic recombination. EMBO J. 2007;26:5071–5082. doi: 10.1038/sj.emboj.7601916. PubMed DOI PMC

Gao J, Kim HM, Elia AE, Elledge SJ, Colaiacovo MP. NatB domain-containing CRA-1 antagonizes hydrolase ACER-1 linking acetyl-CoA metabolism to the initiation of recombination during C. elegans meiosis. PLoS Genet. 2015;11:e1005029. doi: 10.1371/journal.pgen.1005029. PubMed DOI PMC

Mets DG, Meyer BJ. Condensins regulate meiotic DNA break distribution, thus crossover frequency, by controlling chromosome structure. Cell. 2009;139:73–86. doi: 10.1016/j.cell.2009.07.035. PubMed DOI PMC

Hayashi M, Chin GM, Villeneuve AM. C. elegans germ cells switch between distinct modes of double-strand break repair during meiotic prophase progression. PLoS Genet. 2007;3:e191. doi: 10.1371/journal.pgen.0030191. PubMed DOI PMC

Reichman R, Shi Z, Malone R, Smolikove S. Mitotic and meiotic functions for the SUMOylation Pathway in the Caenorhabditis elegans germline. Genetics. 2018;208:1421–1441. doi: 10.1534/genetics.118.300787. PubMed DOI PMC

Mateo AR, et al. The p53-like protein CEP-1 is required for meiotic fidelity in C. elegans. Curr. Biol. 2016;26:1148–1158. doi: 10.1016/j.cub.2016.03.036. PubMed DOI PMC

Woglar A, Villeneuve AM. Dynamic architecture of DNA repair complexes and the synaptonemal complex at sites of meiotic recombination. Cell. 2018;173:1678–1691.e1616. doi: 10.1016/j.cell.2018.03.066. PubMed DOI PMC

Reuben M, Lin R. Germline X chromosomes exhibit contrasting patterns of histone H3 methylation in Caenorhabditis elegans. Dev. Biol. 2002;245:71–82. doi: 10.1006/dbio.2002.0634. PubMed DOI

Penkner AM, et al. Meiotic chromosome homology search involves modifications of the nuclear envelope protein Matefin/SUN-1. Cell. 2009;139:920–933. doi: 10.1016/j.cell.2009.10.045. PubMed DOI

Woglar A, et al. Matefin/SUN-1 phosphorylation is part of a surveillance mechanism to coordinate chromosome synapsis and recombination with meiotic progression and chromosome movement. PLoS Genet. 2013;9:e1003335. doi: 10.1371/journal.pgen.1003335. PubMed DOI PMC

Hillers KJ, Villeneuve AM. Analysis of meiotic recombination in Caenorhabditis elegans. Methods Mol. Biol. 2009;557:77–97. doi: 10.1007/978-1-59745-527-5_7. PubMed DOI

Lim JG, Stine RR, Yanowitz JL. Domain-specific regulation of recombination in Caenorhabditis elegans in response to temperature, age and sex. Genetics. 2008;180:715–726. doi: 10.1534/genetics.108.090142. PubMed DOI PMC

Patel CN, Koh DW, Jacobson MK, Oliveira MA. Identification of three critical acidic residues of poly(ADP-ribose) glycohydrolase involved in catalysis: determining the PARG catalytic domain. Biochem. J. 2005;388:493–500. doi: 10.1042/BJ20040942. PubMed DOI PMC

Gupte R, Liu Z, Kraus WL. PARPs and ADP-ribosylation: recent advances linking molecular functions to biological outcomes. Genes Dev. 2017;31:101–126. doi: 10.1101/gad.291518.116. PubMed DOI PMC

Weaver AN, Yang ES. Beyond DNA repair: additional functions of PARP-1 in cancer. Front. Oncol. 2013;3:290. doi: 10.3389/fonc.2013.00290. PubMed DOI PMC

Ray Chaudhuri A, Nussenzweig A. The multifaceted roles of PARP1 in DNA repair and chromatin remodelling. Nat. Rev. Mol. Cell Biol. 2017;18:610–621. doi: 10.1038/nrm.2017.53. PubMed DOI PMC

Gibson BA, Kraus WL. New insights into the molecular and cellular functions of poly(ADP-ribose) and PARPs. Nat. Rev. Mol. Cell Biol. 2012;13:411–424. doi: 10.1038/nrm3376. PubMed DOI

Pasierbek P, et al. A Caenorhabditis elegans cohesion protein with functions in meiotic chromosome pairing and disjunction. Genes Dev. 2001;15:1349–1360. doi: 10.1101/gad.192701. PubMed DOI PMC

Smolikov S, et al. SYP-3 restricts synaptonemal complex assembly to bridge paired chromosome axes during meiosis in Caenorhabditis elegans. Genetics. 2007;176:2015–2025. doi: 10.1534/genetics.107.072413. PubMed DOI PMC

Kleckner N. Chiasma formation: chromatin/axis interplay and the role(s) of the synaptonemal complex. Chromosoma. 2006;115:175–194. doi: 10.1007/s00412-006-0055-7. PubMed DOI

Johzuka K, Ogawa H. Interaction of Mre11 and Rad50: two proteins required for DNA repair and meiosis-specific double-strand break formation in Saccharomyces cerevisiae. Genetics. 1995;139:1521–1532. PubMed PMC

Jagut M, et al. Separable roles for a Caenorhabditis elegans RMI1 homolog in promoting and antagonizing meiotic crossovers ensure faithful chromosome inheritance. PLoS Biol. 2016;14:e1002412. doi: 10.1371/journal.pbio.1002412. PubMed DOI PMC

Saito TT, Youds JL, Boulton SJ, Colaiacovo MP. Caenorhabditis elegans HIM-18/SLX-4 interacts with SLX-1 and XPF-1 and maintains genomic integrity in the germline by processing recombination intermediates. PLoS Genet. 2009;5:e1000735. doi: 10.1371/journal.pgen.1000735. PubMed DOI PMC

Saito TT, Lui DY, Kim HM, Meyer K, Colaiacovo MP. Interplay between structure-specific endonucleases for crossover control during Caenorhabditis elegans meiosis. PLoS Genet. 2013;9:e1003586. doi: 10.1371/journal.pgen.1003586. PubMed DOI PMC

Link J, et al. Transient and partial nuclear lamina disruption promotes chromosome movement in early meiotic prophase. Dev. Cell. 2018;45:212–225.e217. doi: 10.1016/j.devcel.2018.03.018. PubMed DOI PMC

Davis MW, et al. Rapid single nucleotide polymorphism mapping in C. elegans. BMC Genom. 2005;6:118. doi: 10.1186/1471-2164-6-118. PubMed DOI PMC

Loss of Meiotic Double Strand Breaks Triggers Recruitment of Recombination-independent Pro-crossover Factors in C. elegans Spermatogenesis

Overlapping and separable activities of BRA-2 and HIM-17 promote occurrence and regulation of pairing and synapsis during Caenorhabditis elegans meiosis

Genetic and physical interactions reveal overlapping and distinct contributions to meiotic double-strand break formation in C. elegans

PARG and BRCA1-BARD1 cooperative function regulates DNA repair pathway choice during gametogenesis

Phosphorylation of HORMA-domain protein HTP-3 at Serine 285 is dispensable for crossover formation