Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Crossover patterning through condensation and coarsening of pro-crossover factors

Nature Cell Biology volume 27, pages 1161–1174 (2025)Cite this article

Subjects

Abstract

Meiotic recombination mixes genetic information from parental genomes, creating unique combinations of alleles. During meiotic prophase, each homologue pair must undergo at least one crossover to segregate faithfully. Only a few recombination intermediates become crossovers, and these are widely spaced or limited to one per chromosome pair. Mechanisms that regulate crossover number and spacing remain poorly understood. Here we show that, in Caenorhabditis elegans, ‘recombination nodules’, protein assemblies that stabilize recombination intermediates and promote crossover formation, assemble in part through biomolecular condensation and are stabilized by CDK-2 kinase activity. We further demonstrate that essential components of these nodules move along the synaptonemal complex (SC) and do not freely exchange between SCs in the same nucleus. Our findings reveal that recombination nodules behave as active droplets and support a model in which coarsening of these droplets via protein translocation along liquid crystalline SCs underlies crossover patterning.

Access through your institution
Buy or subscribe

This is a preview of subscription content, access via your institution

Access options

Access through your institution

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Dynamics of SC components and pro-CO proteins.
Fig. 2: Compartmentalization of CO signals by individual SCs.
Fig. 3: ZHP-3 undergoes liquid–liquid phase separation in cultured human cells and in vitro.
Fig. 4: CDK-2 interacts with COSA-1 and is required for CO formation.
Fig. 5: CDK-2 is required to stabilize RNs.
Fig. 6: CDK-2 activity is essential for RN stability and CO formation.
Fig. 7: Models and simulations of RN formation and coarsening.

Similar content being viewed by others

Data availability

The mass spectrometry results have been deposited to the ProteomeXchange Consortium via the iProX repository, with the dataset identifier IPX0010864000 (ProteomeXchange ID: PXD059805). Predicted ZHP-3 and ZHP-4 proteins from Caenorhabditis species that were reanalysed here are available from the Caenorhabditis Genomes Project (http://download.caenorhabditis.org) under accession codes (ZHP-3 orthologues: CELEG.K02B12.8b.1, CINOP.Sp34_10315400.t1, CSP54.g5162.t1, CSP51.g11866.t1, CSP41.g23732.t1, CSP26.g12404.t1, CWALL.g12289.t1, CSP33.g12881.t1, CDOUG.g14799.t1, CSP33.g12881.t2, CSINI.Csp5_scaffold_00204.g6990.t1, CNIGO.Cni-zhp-3, CBRIG.CBG12523.1, CDOUG.g14799.t2, CSP44.g24455.t1, and ZHP-4 orthologs: CELEG.Y39B6A.16.1, CSP54.g8031.t1, CINOP.Sp34_50420510.t1, CSP40.g12044.t1, CSP48.g22458.t1, CSP33.g213.t1, CWALL.g558.t1, CSP41.g17433.t2, CTROP.Csp11.Scaffold630.g17624.t1, CSP29.g19200.t1, CDOUG.g2285.t1, CSP32.g3210.t1 and CBRIG.CBG24873.1). The C. elegans proteomic database used for mass spectrometric analysis was downloaded from UniProt (https://www.uniprot.org). The cdk-2 exon sequences are available at WormBase (https://wormbase.org) under the gene CELE_K03E5.3. Source data are provided with this paper. All other data supporting the findings of this study are available from the corresponding authors on reasonable request.

Code availability

R codes for generating corresponding graphs are provided in the Supplementary Information.

References

  1. Mercier, R., Mézard, C., Jenczewski, E., Macaisne, N. & Grelon, M. The molecular biology of meiosis in plants. Annu. Rev. Plant Biol. 66, 297–327 (2015).

    Article  CAS  PubMed  Google Scholar 

  2. Muller, H. J. The mechanism of crossing-over. Am. Nat. 50, 193–221 (1916).

    Article  Google Scholar 

  3. Sturtevant, A. H. The behavior of the chromosomes as studied through linkage. Z. Indukt. Abstamm. Vererbungslehre 13, 234–287 (1915).

    Google Scholar 

  4. Carpenter, A. T. Electron microscopy of meiosis in Drosophila melanogaster females: II. The recombination nodule–a recombination-associated structure at pachytene? Proc. Natl Acad. Sci. USA 72, 3186–3189 (1975).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Carpenter, A. T. Synaptonemal complex and recombination nodules in wild-type Drosophila melanogaster females. Genetics 92, 511–541 (1979).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Carpenter, A. T. Recombination nodules and the mechanism of crossing-over in Drosophila. Symp. Soc. Exp. Biol. 38, 233–243 (1984).

    CAS  PubMed  Google Scholar 

  7. von Wettstein, D., Rasmussen, S. W. & Holm, P. B. The synaptonemal complex in genetic segregation. Annu. Rev. Genet. 18, 331–411 (1984).

    Article  Google Scholar 

  8. Anderson, L. K. & Stack, S. M. Recombination nodules in plants. Cytogenet. Genome Res. 109, 198–204 (2005).

    Article  CAS  PubMed  Google Scholar 

  9. Pazhayam, N. M., Turcotte, C. A. & Sekelsky, J. Meiotic crossover patterning. Front. Cell Dev. Biol. 9, 1940 (2021).

    Article  Google Scholar 

  10. Colaiácovo, M. P. et al. Synaptonemal complex assembly in C. elegans is dispensable for loading strand-exchange proteins but critical for proper completion of recombination. Dev. Cell 5, 463–474 (2003).

    Article  PubMed  Google Scholar 

  11. MacQueen, A. J. Synapsis-dependent and -independent mechanisms stabilize homolog pairing during meiotic prophase in C. elegans. Genes Dev. 16, 2428–2442 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Kelly, K. O., Dernburg, A. F., Stanfield, G. M. & Villeneuve, A. M. Caenorhabditis elegans msh-5 is required for both normal and radiation-induced meiotic crossing over but not for completion of meiosis. Genetics 156, 617–630 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Agarwal, S. & Roeder, G. S. Zip3 provides a link between recombination enzymes and synaptonemal complex proteins. Cell 102, 245–255 (2000).

    Article  CAS  PubMed  Google Scholar 

  14. Chelysheva, L. et al. The Arabidopsis HEI10 is a new ZMM protein related to Zip3. PLoS Genet. 8, e1002799 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Nguyen, H., Labella, S., Silva, N., Jantsch, V. & Zetka, M. C. elegans ZHP-4 is required at multiple distinct steps in the formation of crossovers and their transition to segregation competent chiasmata. PLoS Genet. 14, e1007776 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  16. Zhang, L., Köhler, S., Rillo-Bohn, R. & Dernburg, A. F. A compartmentalized signaling network mediates crossover control in meiosis. eLife 7, e30789 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  17. De Muyt, A. et al. E3 ligase Hei10: a multifaceted structure-based signaling molecule with roles within and beyond meiosis. Genes Dev. 28, 1111–1123 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  18. Reynolds, A. et al. RNF212 is a dosage-sensitive regulator of crossing-over during mammalian meiosis. Nat. Genet. 45, 269–278 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Qiao, H. et al. Antagonistic roles of ubiquitin ligase HEI10 and SUMO ligase RNF212 regulate meiotic recombination. Nat. Genet. 46, 194–199 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Condezo, Y. B. et al. RNF212B E3 ligase is essential for crossover designation and maturation during male and female meiosis in the mouse. Proc. Natl Acad. Sci. USA 121, e2320995121 (2024).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Yokoo, R. et al. COSA-1 reveals robust homeostasis and separable licensing and reinforcement steps governing meiotic crossovers. Cell 149, 75–87 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Holloway, J. K., Sun, X., Yokoo, R., Villeneuve, A. M. & Cohen, P. E. Mammalian CNTD1 is critical for meiotic crossover maturation and deselection of excess precrossover sites. J. Cell Biol. 205, 633–641 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Ashley, T., Walpita, D. & de Rooij, D. G. Localization of two mammalian cyclin dependent kinases during mammalian meiosis. J. Cell Sci. 114, 685–693 (2001).

    Article  CAS  PubMed  Google Scholar 

  24. Haversat, J. et al. Robust designation of meiotic crossover sites by CDK-2 through phosphorylation of the MutSγ complex. Proc. Natl Acad. Sci. USA 119, e2117865119 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Jaramillo-Lambert, A., Ellefson, M., Villeneuve, A. M. & Engebrecht, J. Differential timing of S phases, X chromosome replication, and meiotic prophase in the C. elegans germ line. Dev. Biol. 308, 206–221 (2007).

    Article  CAS  PubMed  Google Scholar 

  26. Woglar, A. & Villeneuve, A. M. Dynamic architecture of DNA repair complexes and the synaptonemal complex at sites of meiotic recombination. Cell 173, 1678–1691 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Bhalla, N., Wynne, D. J., Jantsch, V. & Dernburg, A. F. ZHP-3 acts at crossovers to couple meiotic recombination with synaptonemal complex disassembly and bivalent formation in C. elegans. PLoS Genet. 4, e1000235 (2008).

    Article  PubMed  PubMed Central  Google Scholar 

  28. 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. 24, 7998–8006 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Rog, O., Köhler, S. & Dernburg, A. F. The synaptonemal complex has liquid crystalline properties and spatially regulates meiotic recombination factors. eLife 6, e21455 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  30. Durand, S. et al. Joint control of meiotic crossover patterning by the synaptonemal complex and HEI10 dosage. Nat. Commun. 13, 5999 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Fozard, J. A., Morgan, C. & Howard, M. Coarsening dynamics can explain meiotic crossover patterning in both the presence and absence of the synaptonemal complex. eLife 12, e79408 (2023).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Morgan, C. et al. Diffusion-mediated HEI10 coarsening can explain meiotic crossover positioning in Arabidopsis. Nat. Commun. 12, 4674 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Hinde, E. et al. The impact of mitotic versus interphase chromatin architecture on the molecular flow of EGFP by pair correlation analysis. Biophys. J. 100, 1829–1836 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Ries, J., Chiantia, S. & Schwille, P. Accurate determination of membrane dynamics with line-scan FCS. Biophys. J. 96, 1999–2008 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Digman, M. A. & Gratton, E. Lessons in fluctuation correlation spectroscopy. Annu. Rev. Phys. Chem. 62, 645–668 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Day, C. A. & Kang, M. The utility of fluorescence recovery after photobleaching (FRAP) to study the plasma membrane. Membranes 13, 492 (2023).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Harwardt, M.-L. I. E., Dietz, M. S., Heilemann, M. & Wohland, T. SPT and imaging FCS provide complementary information on the dynamics of plasma membrane molecules. Biophys. J. 114, 2432–2443 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Digman, M. A. & Gratton, E. Imaging barriers to diffusion by pair correlation functions. Biophys. J. 97, 665–673 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Miné-Hattab, J. & Rothstein, R. Increased chromosome mobility facilitates homology search during recombination. Nat. Cell Biol. 14, 510–517 (2012).

    Article  PubMed  Google Scholar 

  40. Kim, Y. et al. The chromosome axis controls meiotic events through a hierarchical assembly of HORMA domain proteins. Dev. Cell 31, 487–502 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. von Diezmann, L. & Rog, O. Single-molecule tracking of chromatin-associated proteins in the C. elegans gonad. J. Phys. Chem. B 125, 6162–6170 (2021).

    Article  Google Scholar 

  42. Pattabiraman, D., Roelens, B., Woglar, A. & Villeneuve, A. M. Meiotic recombination modulates the structure and dynamics of the synaptonemal complex during C. elegans meiosis. PLoS Genet. 13, e1006670 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  43. Nadarajan, S. et al. The MAP kinase pathway coordinates crossover designation with disassembly of synaptonemal complex proteins during meiosis. eLife 5, e12039 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  44. 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. 9, e1003674 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Von Diezmann, L., Bristow, C. & Rog, O. Diffusion within the synaptonemal complex can account for signal transduction along meiotic chromosomes. Mol. Biol. Cell 35, ar148 (2024).

    Article  Google Scholar 

  46. Hillers, K. J. & Villeneuve, A. M. Chromosome-wide control of meiotic crossing over in C. elegans. Curr. Biol. 13, 1641–1647 (2003).

    Article  CAS  PubMed  Google Scholar 

  47. Yoshimura, J. et al. Recompleting the Caenorhabditis elegans genome. Genome Res. 29, 1009–1022 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Abramson, J. et al. Accurate structure prediction of biomolecular interactions with AlphaFold 3. Nature 630, 493–500 (2024).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. de la Cova, C. C., Townley, R. & Greenwald, I. Negative feedback by conserved kinases patterns the degradation of Caenorhabditis elegans Raf in vulval fate patterning. Development 147, dev195941 (2020).

    Article  PubMed  PubMed Central  Google Scholar 

  50. Viera, A. et al. CDK2 is required for proper homologous pairing, recombination and sex-body formation during male mouse meiosis. J. Cell Sci. 122, 2149–2159 (2009).

    Article  CAS  PubMed  Google Scholar 

  51. Kim, H. J., Liu, C., Zhang, L. & Dernburg, A. F. MJL-1 is a nuclear envelope protein required for homologous chromosome pairing and regulation of synapsis during meiosis in C. elegans. Sci. Adv. 9, eadd1453 (2023).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Sato, A. et al. Cytoskeletal forces span the nuclear envelope to coordinate meiotic chromosome pairing and synapsis. Cell 139, 907–919 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Lopez, M. S., Kliegman, J. I. & Shokat, K. M. in Methods in Enzymology vol. 548 (ed Shokat, K. M.) 189–213 (Elsevier, 2014).

  54. Moses, A. M., Hériché, J.-K. & Durbin, R. Clustering of phosphorylation site recognition motifs can be exploited to predict the targets of cyclin-dependent kinase. Genome Biol. 8, R23 (2007).

    Article  PubMed  PubMed Central  Google Scholar 

  55. Kirschbaum, J. & Zwicker, D. Controlling biomolecular condensates via chemical reactions. J. R. Soc. Interface 18, 20210255 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Söding, J., Zwicker, D., Sohrabi-Jahromi, S., Boehning, M. & Kirschbaum, J. Mechanisms for active regulation of biomolecular condensates. Trends Cell Biol. 30, 4–14 (2020).

    Article  PubMed  Google Scholar 

  57. Schmekel, K. & Daneholt, B. Evidence for close contact between recombination nodules and the central element of the synaptonemal complex. Chromosome Res. 6, 155–159 (1998).

    Article  CAS  PubMed  Google Scholar 

  58. Zickler, D. Development of the synaptonemal complex and the “recombination nodules” during meiotic prophase in the seven bivalents of the fungus Sordaria macrospora Auersw. Chromosoma 61, 289–316 (1977).

    Article  CAS  PubMed  Google Scholar 

  59. Banani, S. F. et al. Compositional control of phase-separated cellular bodies. Cell 166, 651–663 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Wheeler, R. J. & Hyman, A. A. Controlling compartmentalization by non-membrane-bound organelles. Philos. Trans. R. Soc. B 373, 20170193 (2018).

    Article  Google Scholar 

  61. Hyman, A. A., Weber, C. A. & Jülicher, F. Liquid–liquid phase separation in biology. Annu. Rev. Cell Dev. Biol. 30, 39–58 (2014).

    Article  CAS  PubMed  Google Scholar 

  62. Weber, C. A., Zwicker, D., Jülicher, F. & Lee, C. F. Physics of active emulsions. Rep. Prog. Phys. 82, 064601 (2019).

    Article  CAS  PubMed  Google Scholar 

  63. Zwicker, D., Hyman, A. A. & Jülicher, F. Suppression of Ostwald ripening in active emulsions. Phys. Rev. E 92, 012317 (2015).

    Article  Google Scholar 

  64. Stauffer, W. T., Zhang, L. & Dernburg, A. in Biophysics, Biology and Biophotonics IV: The Crossroads (eds. Wax, A. & Backman, V.) 8 (SPIE, 2019); https://doi.org/10.1117/12.2513378

  65. Otto, S. P. & Payseur, B. A. Crossover interference: shedding light on the evolution of recombination. Annu. Rev. Genet. 53, 19–44 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Rafiei, N. & Ronceret, A. Crossover interference mechanism: new lessons from plants. Front. Cell Dev. Biol. 11, 1156766 (2023).

    Article  PubMed  PubMed Central  Google Scholar 

  67. Zickler, D. & Kleckner, N. Meiosis: dances between homologs. Annu. Rev. Genet. 57, 1–63 (2023).

    Article  CAS  PubMed  Google Scholar 

  68. Lloyd, A. Crossover patterning in plants. Plant Reprod. 36, 55–72 (2023).

    Article  PubMed  Google Scholar 

  69. Holliday, R. Recombination and meiosis. Philos. Trans. R. Soc. Lond. B 277, 359–370 (1977).

    Article  CAS  Google Scholar 

  70. Goodyer, W. et al. HTP-3 links DSB formation with homolog pairing and crossing over during C. elegans meiosis. Dev. Cell 14, 263–274 (2008).

    Article  CAS  PubMed  Google Scholar 

  71. Kohli, J. & Bähler, J. Homologous recombination in fission yeast: absence of crossover interference and synaptonemal complex. Experientia 50, 295–306 (1994).

    Article  CAS  PubMed  Google Scholar 

  72. Lukaszewicz, A., Howard-Till, R. A. & Loidl, J. Mus81 nuclease and Sgs1 helicase are essential for meiotic recombination in a protist lacking a synaptonemal complex. Nucleic Acids Res. 41, 9296–9309 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  73. Libuda, D. E., Uzawa, S., Meyer, B. J. & Villeneuve, A. M. Meiotic chromosome structures constrain and respond to designation of crossover sites. Nature 502, 703–706 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  74. Gordon, S. G., Kursel, L. E., Xu, K. & Rog, O. Synaptonemal complex dimerization regulates chromosome alignment and crossover patterning in meiosis. PLoS Genet. 17, e1009205 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  75. Köhler, S., Wojcik, M., Xu, K. & Dernburg, A. F. Dynamic molecular architecture of the synaptonemal complex. Sci. Adv. 11, eadq9374 (2025).

    Article  PubMed  PubMed Central  Google Scholar 

  76. Capilla-Pérez, L. et al. The synaptonemal complex imposes crossover interference and heterochiasmy in Arabidopsis. Proc. Natl Acad. Sci. USA 118, e2023613118 (2021).

    Article  PubMed  PubMed Central  Google Scholar 

  77. France, M. G. et al. ZYP1 is required for obligate cross-over formation and cross-over interference in Arabidopsis. Proc. Natl Acad. Sci. USA 118, e2021671118 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  78. Zalevsky, J., MacQueen, A. J., Duffy, J. B., Kemphues, K. J. & Villeneuve, A. M. Crossing over during Caenorhabditis elegans meiosis requires a conserved MutS-based pathway that is partially dispensable in budding yeast. Genetics 153, 1271–1283 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  79. Kneitz, B. et al. MutS homolog 4 localization to meiotic chromosomes is required for chromosome pairing during meiosis in male and female mice. Genes Dev. 14, 1085–1097 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  80. Yu, Z., Kim, Y. & Dernburg, A. F. Meiotic recombination and the crossover assurance checkpoint in Caenorhabditis elegans. Semin. Cell Dev. Biol. 54, 106–116 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  81. Hinch, A. G. et al. Factors influencing meiotic recombination revealed by whole-genome sequencing of single sperm. Science 363, eaau8861 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  82. Zhang, L., Ward, J. D., Cheng, Z. & Dernburg, A. F. The auxin-inducible degradation (AID) system enables versatile conditional protein depletion in C. elegans. Development 142, 4374–4384 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  83. Ooi, S. L., Priess, J. R. & Henikoff, S. Histone H3.3 variant dynamics in the germline of Caenorhabditis elegans. PLoS Genet. 2, e97 (2006).

    Article  PubMed  PubMed Central  Google Scholar 

  84. Janisiw, E., Dello Stritto, M. R., 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. 14, e1007653 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  85. Cowan, C. R. & Hyman, A. A. Cyclin E–Cdk2 temporally regulates centrosome assembly and establishment of polarity in Caenorhabditis elegans embryos. Nat. Cell Biol. 8, 1441–1447 (2006).

    Article  CAS  PubMed  Google Scholar 

  86. Rog, O. & Dernburg, A. F. Direct visualization reveals kinetics of meiotic chromosome synapsis. Cell Rep. 10, 1639–1645 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  87. Woglar, A. et al. Quantitative cytogenetics reveals molecular stoichiometry and longitudinal organization of meiotic chromosome axes and loops. PLoS Biol. 18, e3000817 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  88. Sarov, M. et al. A genome-scale resource for in vivo tag-based protein function exploration in C. elegans. Cell 150, 855–866 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  89. Zhang, L. et al. Recruitment of Polo-like kinase couples synapsis to meiotic progression via inactivation of CHK-2. eLife 12, e84492 (2023).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  90. Hollis, J. A. et al. Excess crossovers impede faithful meiotic chromosome segregation in C. elegans. PLoS Genet. 16, e1009001 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  91. Wachsmuth, M., Waldeck, W. & Langowski, J. Anomalous diffusion of fluorescent probes inside living cell nuclei investigated by spatially-resolved fluorescence correlation spectroscopy. J. Mol. Biol. 298, 677–689 (2000).

    Article  CAS  PubMed  Google Scholar 

  92. Qian, H. & Elson, E. L. Analysis of confocal laser-microscope optics for 3-D fluorescence correlation spectroscopy. Appl. Opt. 30, 1185–1195 (1991).

    Article  CAS  PubMed  Google Scholar 

  93. Vukojević, V. et al. Study of molecular events in cells by fluorescence correlation spectroscopy. Cell. Mol. Life Sci. 62, 535–550 (2005).

    Article  PubMed  PubMed Central  Google Scholar 

  94. Harper, N. C. et al. Pairing centers recruit a polo-like kinase to orchestrate meiotic chromosome dynamics in C. elegans. Dev. Cell 21, 934–947 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  95. MacQueen, A. J. et al. Chromosome sites play dual roles to establish homologous synapsis during meiosis in C. elegans. Cell 123, 1037–1050 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  96. Phillips, C. M. et al. HIM-8 binds to the X chromosome pairing center and mediates chromosome-specific meiotic synapsis. Cell 123, 1051–1063 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  97. Song, X. et al. Phase separation of EB1 guides microtubule plus-end dynamics. Nat. Cell Biol. 25, 79–91 (2023).

    Article  CAS  PubMed  Google Scholar 

  98. Fu, W. et al. Multivalent interactions of Septin 6 promote the establishment of epithelial cell polarity. J. Mol. Cell Biol. https://doi.org/10.1093/jmcb/mjaf003 (2025).

  99. Sturm, Á., Saskői, É., Tibor, K., Weinhardt, N. & Vellai, T. Highly efficient RNAi and Cas9-based auto-cloning systems for C. elegans research. Nucleic Acids Res. 46, e105–e105 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  100. Notredame, C., Higgins, D. G. & Heringa, J. T-coffee: a novel method for fast and accurate multiple sequence alignment. J. Mol. Biol. 302, 205–217 (2000).

    Article  CAS  PubMed  Google Scholar 

  101. Jumper, J. et al. Highly accurate protein structure prediction with AlphaFold. Nature 596, 583–589 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  102. Yang, Y. et al. COSA-1 mediated pro-crossover complex formation promotes meiotic crossing over in C. elegans. Nucleic Acids Res. 52, 4375–4392 (2024).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This Article is dedicated to the memory of Adelaide Carpenter (1944–2024), whose insights into the physical nature of recombination nodules led to the work presented here. Adelaide also provided encouragement and helpful comments on an early version of this manuscript. We thank J. S. Wang, J. D. Robinson and F. Xiong for experimental assistance, N. Silva for sharing the gfp::msh-5 strain, D. Libuda for sharing the gfp::cosa-1; meT7 strain and D. Updike, J. Nance and S. Smolikove for sharing split-GFP strains. This work was partially supported by grants 2022YFA1303100 and 32090040 (to X.Y.). We thank members of the Dernburg Lab and G. Karpen for helpful discussions and comments on the manuscript.

Author information

Author notes

  1. These authors contributed equally: Liangyu Zhang, Weston Stauffer, Chenshu Liu, Hengyi Shao.

Authors and Affiliations

  1. Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, USA

    Liangyu Zhang, Weston Stauffer, Chenshu Liu, Hengyi Shao, Noor Abuzahriyeh, Rui Jiang & Abby F. Dernburg

  2. Howard Hughes Medical Institute, Chevy Chase, MD, USA

    Liangyu Zhang, Weston Stauffer, Chenshu Liu & Abby F. Dernburg

  3. Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA

    Liangyu Zhang & Abby F. Dernburg

  4. California Institute for Quantitative Biosciences, Berkeley, CA, USA

    Liangyu Zhang & Abby F. Dernburg

  5. MOE Key Laboratory for Membraneless Organelles and Cellular Dynamics, Hefei National Research Center for Physical Sciences at the Microscale, University of Science and Technology of China, Hefei, China

    Liangyu Zhang, Hengyi Shao, Xing Liu & Xuebiao Yao

  6. Max Planck Institute for Dynamics and Self-Organization, Göttingen, Germany

    David Zwicker

Authors
  1. Liangyu Zhang
    View author publications

    Search author on:PubMed Google Scholar

  2. Weston Stauffer
    View author publications

    Search author on:PubMed Google Scholar

  3. Chenshu Liu
    View author publications

    Search author on:PubMed Google Scholar

  4. Hengyi Shao
    View author publications

    Search author on:PubMed Google Scholar

  5. Noor Abuzahriyeh
    View author publications

    Search author on:PubMed Google Scholar

  6. Rui Jiang
    View author publications

    Search author on:PubMed Google Scholar

  7. David Zwicker
    View author publications

    Search author on:PubMed Google Scholar

  8. Xing Liu
    View author publications

    Search author on:PubMed Google Scholar

  9. Xuebiao Yao
    View author publications

    Search author on:PubMed Google Scholar

  10. Abby F. Dernburg
    View author publications

    Search author on:PubMed Google Scholar

Contributions

Conceptualization: L.Z. and A.F.D.; methodology: L.Z., W.S., C.L., H.S., D.Z., X.L., X.Y. and A.F.D.; investigation: L.Z., W.S., C.L., H.S., N.A. and R.J.; visualization: L.Z., W.S., C.L., H.S., D.Z. and A.F.D.; funding acquisition: X.Y. and A.F.D.; project administration: L.Z. and A.F.D.; supervision: L.Z., X.Y. and A.F.D.; writing—original draft: L.Z., W.S., D.Z. and A.F.D.; writing—review and editing: all authors.

Corresponding authors

Correspondence to Liangyu Zhang, Xuebiao Yao or Abby F. Dernburg.

Ethics declarations

Competing interests

The authors declare no competing interests.

Peer review

Peer review information

Nature Cell Biology thanks Denes Hnisz, Akira Shinohara and the other, anonymous, reviewer(s) for their contribution to the peer review of this work. Peer reviewer reports are available.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Extended Data Fig. 1 Dynamics of SC components and the pro-CO proteins.

a, Diffusion coefficients derived from autocorrelation functions (ACFs) for individual voxels in confocal line scans for each protein. Black boxes and bars indicate the average and median values, respectively. Number of voxels: n = 152 (HIS-72::GFP, LP), 104 (HTP-3::GFP, LP), 80 (GFP::HIM-3, LP), 104 (SYP-1::GFP, LP), 56 (SYP-2::GFP, LP), 112 (GFP::SYP-3, EP), 120 (GFP::SYP-3, LP), 104 (ZHP-3::GFP, SC, mid-pachytene), and 112 (ZHP-3::GFP, LRN, late pachytene), respectively, pooled from two independent replicates. Voxels are from the line scans in Fig. 1c. See Fig. 1c for the number of scans for each genotype or condition. One scan per SC. Two-tailed Mann-Whitney U test. Detailed statistical comparisons (including P-values) are reported in Supplementary Table 2 (red). SC, synaptonemal complex, EP, early pachytene. LP, late pachytene. LRN, late recombination nodule. Protein names are colorized based on the structures that they associate with (chromatin, chromosome axis, SC, and/or RNs). b, Movement indices based on paired correlation functions (pCFs) of single voxel pairs. Black boxes and bars indicate the average and median, respectively. Number of voxel pairs: n = 91 (SYP-1::GFP, LP), 49 (SYP-2::GFP, LP), 105 (GFP::SYP-3, EP), 105 (GFP::SYP-3, LP), 91 (ZHP-3::GFP, SC, mid-pachytene), and 91 (ZHP-3::GFP, LRN, late pachytene), respectively, pooled from two independent replicates. Voxel pairs are from the line scans in Fig. 1d. See Fig. 1d for the number of scans for each genotype or condition. One scan per SC. Two-tailed Mann-Whitney U test. Detailed statistical comparisons (including P-values) are reported in Supplementary Table 3 (red). c, Representative FRAP analysis of ZHP-3::GFP along SCs in a mid-pachytene nucleus. The green arrow indicates the bleached region. ZHP-3::GFP intensity recovered within minutes after photobleaching, while HTP-3 (see Extended Data Fig. 2, performed in parallel) showed no recovery. Elapsed time is expressed in minutes:seconds relative to the moment of bleaching. Scale bar, 5 µm. d, FRAP quantification. The average intensities of the regions of interest over time were normalized against their average intensity before photobleaching. Means ± s.e.m. were plotted. n = 12 nuclei from 7 animals for each condition, pooled from two independent replicates. For each nucleus, a bleached region and a similar unbleached (control) region in the same nucleus were measured. Source numerical data are available in source data.

Source data

Extended Data Fig. 2 SC confines the movement of its components within individual SCs.

a, Representative time-lapse images showing fluorescence recovery after photobleaching (FRAP) of GFP::SYP-3 along a segment of an SC in a mid-pachytene nucleus. The green arrow indicates the photobleached region. Time is indicated in minutes:seconds relative to the moment of bleaching. b, Quantification of GFP::SYP-3 intensity over time as described in panel a. In each nucleus, a bleached region and a similar unbleached control region were measured. Intensities were normalized based on prebleached values in the region of interest. Time is indicated in minutes after bleaching. n = 15 nuclei from 6 animals for each condition, pooled from two independent replicates. c, Representative time-lapse images showing FRAP of GFP::SYP-3 following bleaching of an entire SC in a mid-pachytene nucleus. The green box indicates the photobleached region. d, Quantification of GFP::SYP-3 intensity over time as described in c. n = 14 nuclei from 6 animals for each condition, pooled from two independent replicates. e, Representative images showing FRAP of HTP-3::GFP along a segment of paired chromosome axes in a mid-pachytene nucleus. The green arrow indicates the photobleached region. f, Quantification of HTP-3::GFP intensity over time as described in panel e. n = 12 nuclei from 5 animals for each condition, pooled from two independent replicates. g, FRAP of HTP-3::GFP on an entire chromosome pair in a mid-pachytene nucleus. The green box indicates the photobleached region. h, Quantification of HTP-3::GFP intensity over time as described in panel g. n = 12 nuclei from 5 animals for each condition, pooled from two independent replicates. Scale bars in a, c, e and g: 5 μm. Means ± s.e.m. were plotted in b, d, f, h. Source numerical data are available in source data.

Source data

Extended Data Fig. 3 LRNs in meT7 nuclei.

a, Schematic of a late pachytene nucleus in a hermaphrodite homozygous for the III; X; IV triple fusion chromosome meT7. Paired axes are shown in magenta to correspond to HTP-3 immunofluorescence shown in panel b. The meT7 chromosome is marked by HIM-8 (red), which binds the pairing center region on the normal X chromosomes. Late recombination nodules (LRNs) are marked by COSA-1. b, Representative immunofluorescence images showing late pachytene nuclei with paired meT7 fusion chromosomes. GFP::COSA-1 intensity at LRNs within these nuclei is heterogeneous, in contrast to wild-type oocytes. Scale bar, 5 µm. c-f, Quantification of GFP::COSA-1 and ZHP-3::GFP fluorescence intensities at individual LRNs in meT7 nuclei at late pachytene. Each dot represents the summed intensity for one focus. Lines connect values for foci from the same nucleus. The foci associated with meT7 foci were identified by HIM-8 and HTP-3 staining. The ‘non-meT7’ foci are on the normal autosomes, shown in an arbitrary order (these chromosomes were not identified individually). Intensities were normalized such that the average value of foci on non-meT7 chromosomes was 1. For nuclei with 5 COSA-1 or ZHP-3 foci, the brighter foci on meT7 were mandatorily assigned to the first column. n = 46, 40, 53, and 44 nuclei, from 6, 6, 16, and 16 animals, for nuclei with 4 GFP::COSA-1 foci, nuclei with 5 GFP::COSA-1 foci, nuclei with 4 ZHP-3::GFP foci, and nuclei with 5 ZHP-3::GFP foci, respectively, pooled from two independent replicates. See Fig. 2 for detailed comparisons. Source numerical data are available in source data.

Source data

Extended Data Fig. 4 Properties of pro-CO proteins expressed in HEK293T cells.

a, AlphaFold3 predictions illustrating the structural features of ZHP-3 and ZHP-4 heterotetramer, highlighting extensive intrinsically disordered regions in the C-terminal tails of both proteins (upper). a.a., amino acid. b, Left: an image of droplets formed by GFP-ZHP-3 a.a.169-389 in a HEK293T cell. Right: enlarged views displaying time-lapse images of the boxed area on the left, demonstrating droplet fusion. Time is indicated in seconds. Scale bars, 10 μm (left) and 1 μm (right). c, Fluorescence recovery in a droplet of GFP-ZHP-3 a.a.169-389 after photobleaching. GFP-ZHP-3 a.a.169-389 were transiently expressed in HEK293T cells. ‘Pre’ indicates pre-photobleach, and the white square highlights the photobleached area. Time is indicated in seconds. Scale bar, 1 μm. d, Quantitative analysis of the FRAP recovery of ZHP-3 a.a.169-389 as depicted in panel c. Data are presented as mean ± s.d. at each time point. n = 16 droplets from two independent replicates. e, Representative fluorescent microscopy images of live cells showing the full-length (FL) pro-crossover proteins and their truncated variants transiently expressed in HEK293T cells. These images were obtained in parallel with those presented in Fig. 3b. Dashed white lines indicate the cell membranes. Scale bar, 10 μm. f, Fluorescent images of live cells showing the co-condensates formed by two IDRs from ZHP-3 and ZHP-4, or three IDRs from ZHP-3, ZHP-4 and MSH-5. IDRs were transiently co-expressed in HEK293T cells. Dashed white and orange lines indicate cell and nuclear membranes, respectively. a.a., amino acid. Scale bar, 10 μm. Source numerical data are available in source data.

Source data

Extended Data Fig. 5 Sensitivity of ZHP-3–ZHP-4 droplets to the high salt concentrations.

a, Coomassie blue-stained gel exhibiting the purified ZHP-3 and ZHP-4 complex from HEK293T cells. GFP-StrepII-ZHP-3 and His-mCherry-ZHP-4 were transiently coexpressed in HEK293T cells and then purified initially using Ni-NTA agarose beads, followed by a secondary purification step with the Strep-Tactin beads. Approximately 100 ng each of ZHP-3 and ZHP-4 were separated in the lane on the right. b, Representative fluorescence, and Differential Interference Contrast (DIC) images showing condensates formed by 50 nM ZHP-3 (green) and 50 nM ZHP-4 (magenta) complex at various salt and 1, 6-hexanediol concentrations in vitro. Scale bar, 1 μm. c-e, Statistical analyses of ZHP-3–ZHP-4 droplets as described in panel b. c, Droplet number per imaging field. n = 20, 10, 11, 13, 12, 24, 23, 12 imaging fields, for 100 mM, 150 mM, 200 mM, 250 mM, 300 mM NaCl and 5%, 10%, 15% hexanediol, respectively, pooled from two independent replicates. Each imaging field = 2023 μm2. d, Total droplet area per imaging field. n = 20, 10, 11, 13, 12, 24, 23, 12 imaging fields for 100 mM, 150 mM, 200 mM, 250 mM, 300 mM NaCl and 5%, 10%, 15% hexanediol, respectively, pooled from two independent replicates. e, Individual droplet size. Only droplets with a size > 0.1 μm2 were scored due to the resolution limit. n = 2046, 893, 401, 126, 18, 3011, 2772, 1047 droplets for 100 mM, 150 mM, 200 mM, 250 mM, 300 mM NaCl and 5%, 10%, 15% hexanediol, respectively, pooled from two independent replicates. Data are presented as mean ± s.d. for c-e. Ordinary one-way ANOVA, Tukey’s multiple comparisons test with a single pooled variance and each P value adjusted for c and d, and two-tailed Kolmogorov-Smirnov test for e. Source numerical data and unprocessed gel are available in source data.

Source data

Extended Data Fig. 6 Depletion of CDK-2 by the AID system and RNAi, and the dispensability of CDK-2 for homologue pairing, synapsis, and DSB induction.

a, Late prophase nuclei in aid::HA::cdk-2; sun-1p::tir1 hermaphrodites stained with anti-HA antibodies show incomplete depletion of CDK-2 by auxin treatment alone, while effective depletion by RNAi + auxin treatment. Arrowheads indicate the remaining CDK-2 signal. b, Early prophase nuclei in aid::HA::cdk-2; sun-1p::tir1 gonads stained for HIM-8, which marks the pairing center region of the X chromosome, and DNA. The robust pairing of HIM-8 foci was observed following depletion of CDK-2. c, Nuclei at mid-prophase stained for HTP-3 and SYP-1. Colocalization of these proteins reveals the formation of SCs along all axes in controls and hermaphrodites depleted of CDK-2. d, Nuclei at mid-prophase stained for RAD-51 and DNA, showing abundant DSB repair intermediates marked by RAD-51 following depletion of CDK-2. e, Dynamics of RAD-51 foci. Hermaphrodite gonads were divided into six zones of equal length spanning the premeiotic through the late pachytene region, as shown in the diagram. The number of RAD-51 foci in each nucleus is shown for each zone, with mean and s.d. values indicated by bars. The numbers of nuclei counted for zones 1, 2, 3, 4, 5, and 6 were as follows: for controls – 74, 113, 113, 127, 86, and 47, respectively; for CDK-2 depletion – 47, 46, 80, 82, 58, and 33, respectively. Data were derived from 2 animals for each condition, pooled from two independent replicates. More nuclei were present in the controls due to the effects of CDK-2 depletion on germline proliferation. ****P = 8.2210e-6 and 3.8779e-7, respectively. Two-tailed Mann-Whitney test. Scale bars in a-d: 5 µm. Source numerical data are available in source data.

Source data

Extended Data Fig. 7 Incomplete depletion of CDK-2 by RNAi or AID alone.

a, Immunolocalization of GFP::MSH-5, ALFA::COSA-1 and SYP-2 in hermaphrodite germlines. Upper panel: recombination nodules (RNs) are not affected by auxin treatment of a control strain expressing CDK-2 without a degron. cdk-2(RNAi) treatment for 24 hours or exposure to 4 mM auxin for 6 hours strongly reduces but does not abolish RNs. ALFA::COSA-1 localization was more sensitive to CDK-2 depletion than GFP::MSH-5. These samples were placed on RNAi or auxin plates and stained in parallel with those shown in Fig. 5. Scale bar, 5 μm. b, Enlargements of the indicated regions from a. Scale bar, 5 μm.

Extended Data Fig. 8 Phosphorylation sites on ZHP-3.

Ser210 and Ser214 on ZHP-3 were frequently identified as phosphorylation sites by mass spectrometric analysis of GFP-strepII-ZHP-3 IDR expressed and purified from HEK293T cells. Raw data are available at ProteomeXchange Consortium via the iProX repository, with the dataset identifier IPX0010864000.

Extended Data Fig. 9 DAPI-staining bodies and LRN formation in various phosphosite mutants.

a-b, Graphs showing the distribution of nuclei containing varying numbers of DAPI-staining bodies (bivalents and univalents) at diakinesis in various phosphosite mutants at 15°C (a) and 20°C (b). In a, n = 147, 67, 140, 138, 139, and 153 scored nuclei from 39, 21, 38, 36, 37, and 39 animals, for wild type, msh-5 13A, zhp-3 4A, zhp-4 2A, zhp-3 4A; zhp-4 2A, and zhp-3 4A; msh-5 13A; zhp-4 2A, respectively, pooled from two independent replicates. In b, n = 193, 127, 158, 153, 151, and 159 scored nuclei from 39, 31, 36, 34, 35, and 41 animals, for wild type, msh-5 13A, zhp-3 4A, zhp-4 2A, zhp-3 4A; zhp-4 2A, and zhp-3 4A; msh-5 13A; zhp-4 2A, respectively, pooled from two independent replicates. One-way ANOVA and post hoc pairwise comparisons using t-tests with pooled s.d.. P value adjustment method: Holm. Comparisons between each genotype in a, not significant. Triple mutants vs. wild type or msh-5 13A in b, P = 0.0051 or 0.0147. Other comparisons in b, not significant. c, Representative immunofluorescence images showing GFP::COSA-1 staining from mid-pachytene to diplotene onset in various phospho-mutants at 25 °C. SC was marketed by SYP-1 staining. Scale bar, 5 μm. Source numerical data are available in source data.

Source data

Supplementary information

Supplementary Information

Supplementary Figs. 1 and 2.

Reporting Summary

Peer Review File

Supplementary Table 1

Supplementary Tables 1–4: detailed statistics in Fig. 1c,d and Extended Data Fig. 1a,b; Supplementary Table 5: viability and fertility of representative transgenic worm strains; Supplementary Table 6: alleles generated in this study; Supplementary Table 7: crRNAs, repair templates and genotyping primers used in this study.

Supplementary Code 1

R codes for generating graphs shown in Figs. 2j–m and 6d,f and Extended Data Figs. 1d, 2b,d,f,h, 3c–f and 9a,b.

Source data

Source Data Fig. 1

Statistical source data.

Source Data Fig. 2

Statistical source data.

Source Data Fig. 3

Statistical source data.

Source Data Fig. 4

Statistical source data.

Source Data Fig. 4

Unprocessed western blots.

Source Data Fig. 6

Statistical source data.

Source Data Extended Data Fig. 1

Statistical source data.

Source Data Extended Data Fig. 2

Statistical source data.

Source Data Extended Data Fig. 3

Statistical source data.

Source Data Extended Data Fig. 4

Statistical source data.

Source Data Extended Data Fig. 5

Statistical source data.

Source Data Extended Data Fig. 5

Unprocessed gels.

Source Data Extended Data Fig. 6

Statistical source data.

Source Data Extended Data Fig. 9

Statistical source data.

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zhang, L., Stauffer, W., Liu, C. et al. Crossover patterning through condensation and coarsening of pro-crossover factors. Nat Cell Biol 27, 1161–1174 (2025). https://doi.org/10.1038/s41556-025-01688-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue date:

  • DOI: https://doi.org/10.1038/s41556-025-01688-9

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing