Skip to main content
NCBI home page
The Plant Cell logoLink to The Plant Cell
. 1999 May;11(5):809–824. doi: 10.1105/tpc.11.5.809

Three-dimensional microscopy of the Rad51 recombination protein during meiotic prophase.

A E Franklin 1, J McElver 1, I Sunjevaric 1, R Rothstein 1, B Bowen 1, W Z Cande 1
PMCID: PMC144225  PMID: 10330467

Abstract

An open question in meiosis is whether the Rad51 recombination protein functions solely in meiotic recombination or whether it is also involved in the chromosome homology search. To address this question, we have performed three-dimensional high-resolution immunofluorescence microscopy to visualize native Rad51 structures in maize male meiocytes. Maize has two closely related RAD51 genes that are expressed at low levels in differentiated tissues and at higher levels in mitotic and meiotic tissues. Cells and nuclei were specially fixed and embedded in polyacrylamide to maintain both native chromosome structure and the three dimensionality of the specimens. Analysis of Rad51 in maize meiocytes revealed that when chromosomes condense during leptotene, Rad51 is diffuse within the nucleus. Rad51 foci form on the chromosomes at the beginning of zygotene and rise to approximately 500 per nucleus by mid-zygotene when chromosomes are pairing and synapsing. During chromosome pairing, we consistently found two contiguous Rad51 foci on paired chromosomes. These paired foci may identify the sites where DNA sequence homology is being compared. During pachytene, the number of Rad51 foci drops to seven to 22 per nucleus. This higher number corresponds approximately to the number of chiasmata in maize meiosis. These observations are consistent with a role for Rad51 in the homology search phase of chromosome pairing in addition to its known role in meiotic recombination.

Full Text

The Full Text of this article is available as a PDF (837.0 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson L. K., Offenberg H. H., Verkuijlen W. M., Heyting C. RecA-like proteins are components of early meiotic nodules in lily. Proc Natl Acad Sci U S A. 1997 Jun 24;94(13):6868–6873. doi: 10.1073/pnas.94.13.6868. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Aragón-Alcaide L., Reader S., Beven A., Shaw P., Miller T., Moore G. Association of homologous chromosomes during floral development. Curr Biol. 1997 Nov 1;7(11):905–908. doi: 10.1016/s0960-9822(06)00383-6. [DOI] [PubMed] [Google Scholar]
  3. Ashley T., Plug A. W., Xu J., Solari A. J., Reddy G., Golub E. I., Ward D. C. Dynamic changes in Rad51 distribution on chromatin during meiosis in male and female vertebrates. Chromosoma. 1995 Oct;104(1):19–28. doi: 10.1007/BF00352222. [DOI] [PubMed] [Google Scholar]
  4. Barlow A. L., Benson F. E., West S. C., Hultén M. A. Distribution of the Rad51 recombinase in human and mouse spermatocytes. EMBO J. 1997 Sep 1;16(17):5207–5215. doi: 10.1093/emboj/16.17.5207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bass H. W., Marshall W. F., Sedat J. W., Agard D. A., Cande W. Z. Telomeres cluster de novo before the initiation of synapsis: a three-dimensional spatial analysis of telomere positions before and during meiotic prophase. J Cell Biol. 1997 Apr 7;137(1):5–18. doi: 10.1083/jcb.137.1.5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Baumann P., Benson F. E., West S. C. Human Rad51 protein promotes ATP-dependent homologous pairing and strand transfer reactions in vitro. Cell. 1996 Nov 15;87(4):757–766. doi: 10.1016/s0092-8674(00)81394-x. [DOI] [PubMed] [Google Scholar]
  7. Benson F. E., Baumann P., West S. C. Synergistic actions of Rad51 and Rad52 in recombination and DNA repair. Nature. 1998 Jan 22;391(6665):401–404. doi: 10.1038/34937. [DOI] [PubMed] [Google Scholar]
  8. Bishop D. K., Park D., Xu L., Kleckner N. DMC1: a meiosis-specific yeast homolog of E. coli recA required for recombination, synaptonemal complex formation, and cell cycle progression. Cell. 1992 May 1;69(3):439–456. doi: 10.1016/0092-8674(92)90446-j. [DOI] [PubMed] [Google Scholar]
  9. Bishop D. K. RecA homologs Dmc1 and Rad51 interact to form multiple nuclear complexes prior to meiotic chromosome synapsis. Cell. 1994 Dec 16;79(6):1081–1092. doi: 10.1016/0092-8674(94)90038-8. [DOI] [PubMed] [Google Scholar]
  10. Burnham C. R., Stout J. T., Weinheimer W. H., Kowles R. V., Phillips R. L. Chromosome pairing in maize. Genetics. 1972 May;71(1):111–126. doi: 10.1093/genetics/71.1.111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Carpenter A. T. Electron microscopy of meiosis in Drosophila melanogaster females. I. Structure, arrangement, and temporal change of the synaptonemal complex in wild-type. Chromosoma. 1975;51(2):157–182. doi: 10.1007/BF00319833. [DOI] [PubMed] [Google Scholar]
  12. Cheng R., Baker T. I., Cords C. E., Radloff R. J. mei-3, a recombination and repair gene of Neurospora crassa, encodes a RecA-like protein. Mutat Res. 1993 Oct;294(3):223–234. doi: 10.1016/0921-8777(93)90005-2. [DOI] [PubMed] [Google Scholar]
  13. Copenhaver G. P., Browne W. E., Preuss D. Assaying genome-wide recombination and centromere functions with Arabidopsis tetrads. Proc Natl Acad Sci U S A. 1998 Jan 6;95(1):247–252. doi: 10.1073/pnas.95.1.247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Creighton H. B., McClintock B. A Correlation of Cytological and Genetical Crossing-Over in Zea Mays. Proc Natl Acad Sci U S A. 1931 Aug;17(8):492–497. doi: 10.1073/pnas.17.8.492. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Dawe R. K., Sedat J. W., Agard D. A., Cande W. Z. Meiotic chromosome pairing in maize is associated with a novel chromatin organization. Cell. 1994 Mar 11;76(5):901–912. doi: 10.1016/0092-8674(94)90364-6. [DOI] [PubMed] [Google Scholar]
  16. Dernburg A. F., McDonald K., Moulder G., Barstead R., Dresser M., Villeneuve A. M. Meiotic recombination in C. elegans initiates by a conserved mechanism and is dispensable for homologous chromosome synapsis. Cell. 1998 Aug 7;94(3):387–398. doi: 10.1016/s0092-8674(00)81481-6. [DOI] [PubMed] [Google Scholar]
  17. Doebley J., Stec A., Wendel J., Edwards M. Genetic and morphological analysis of a maize-teosinte F2 population: implications for the origin of maize. Proc Natl Acad Sci U S A. 1990 Dec 15;87(24):9888–9892. doi: 10.1073/pnas.87.24.9888. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Dunderdale H. J., Benson F. E., Parsons C. A., Sharples G. J., Lloyd R. G., West S. C. Formation and resolution of recombination intermediates by E. coli RecA and RuvC proteins. Nature. 1991 Dec 19;354(6354):506–510. doi: 10.1038/354506a0. [DOI] [PubMed] [Google Scholar]
  19. Giloh H., Sedat J. W. Fluorescence microscopy: reduced photobleaching of rhodamine and fluorescein protein conjugates by n-propyl gallate. Science. 1982 Sep 24;217(4566):1252–1255. doi: 10.1126/science.7112126. [DOI] [PubMed] [Google Scholar]
  20. Gupta R. C., Bazemore L. R., Golub E. I., Radding C. M. Activities of human recombination protein Rad51. Proc Natl Acad Sci U S A. 1997 Jan 21;94(2):463–468. doi: 10.1073/pnas.94.2.463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hyde H., Davies A. A., Benson F. E., West S. C. Resolution of recombination intermediates by a mammalian activity functionally analogous to Escherichia coli RuvC resolvase. J Biol Chem. 1994 Feb 18;269(7):5202–5209. [PubMed] [Google Scholar]
  22. MOSES M. J. Chromosomal structures in crayfish spermatocytes. J Biophys Biochem Cytol. 1956 Mar 25;2(2):215–218. doi: 10.1083/jcb.2.2.215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Maeshima K., Morimatsu K., Horii T. Purification and characterization of XRad51.1 protein, Xenopus RAD51 homologue: recombinant XRad51.1 promotes strand exchange reaction. Genes Cells. 1996 Dec;1(12):1057–1068. doi: 10.1046/j.1365-2443.1996.d01-224.x. [DOI] [PubMed] [Google Scholar]
  24. Maguire M. P., Riess R. W., Paredes A. M. Evidence from a maize desynaptic mutant points to a probable role of synaptonemal complex central region components in provision for subsequent chiasma maintenance. Genome. 1993 Oct;36(5):797–807. doi: 10.1139/g93-105. [DOI] [PubMed] [Google Scholar]
  25. Maguire M. P., Riess R. W. The relationship of homologous synapsis and crossing over in a maize inversion. Genetics. 1994 May;137(1):281–288. doi: 10.1093/genetics/137.1.281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. McKim K. S., Green-Marroquin B. L., Sekelsky J. J., Chin G., Steinberg C., Khodosh R., Hawley R. S. Meiotic synapsis in the absence of recombination. Science. 1998 Feb 6;279(5352):876–878. doi: 10.1126/science.279.5352.876. [DOI] [PubMed] [Google Scholar]
  27. Moens P. B., Chen D. J., Shen Z., Kolas N., Tarsounas M., Heng H. H., Spyropoulos B. Rad51 immunocytology in rat and mouse spermatocytes and oocytes. Chromosoma. 1997 Sep;106(4):207–215. doi: 10.1007/s004120050241. [DOI] [PubMed] [Google Scholar]
  28. Morita T., Yoshimura Y., Yamamoto A., Murata K., Mori M., Yamamoto H., Matsushiro A. A mouse homolog of the Escherichia coli recA and Saccharomyces cerevisiae RAD51 genes. Proc Natl Acad Sci U S A. 1993 Jul 15;90(14):6577–6580. doi: 10.1073/pnas.90.14.6577. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. New J. H., Sugiyama T., Zaitseva E., Kowalczykowski S. C. Rad52 protein stimulates DNA strand exchange by Rad51 and replication protein A. Nature. 1998 Jan 22;391(6665):407–410. doi: 10.1038/34950. [DOI] [PubMed] [Google Scholar]
  30. Nishinaka T., Shinohara A., Ito Y., Yokoyama S., Shibata T. Base pair switching by interconversion of sugar puckers in DNA extended by proteins of RecA-family: a model for homology search in homologous genetic recombination. Proc Natl Acad Sci U S A. 1998 Sep 15;95(19):11071–11076. doi: 10.1073/pnas.95.19.11071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Padmore R., Cao L., Kleckner N. Temporal comparison of recombination and synaptonemal complex formation during meiosis in S. cerevisiae. Cell. 1991 Sep 20;66(6):1239–1256. doi: 10.1016/0092-8674(91)90046-2. [DOI] [PubMed] [Google Scholar]
  32. Plug A. W., Xu J., Reddy G., Golub E. I., Ashley T. Presynaptic association of Rad51 protein with selected sites in meiotic chromatin. Proc Natl Acad Sci U S A. 1996 Jun 11;93(12):5920–5924. doi: 10.1073/pnas.93.12.5920. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Rockmill B., Sym M., Scherthan H., Roeder G. S. Roles for two RecA homologs in promoting meiotic chromosome synapsis. Genes Dev. 1995 Nov 1;9(21):2684–2695. doi: 10.1101/gad.9.21.2684. [DOI] [PubMed] [Google Scholar]
  34. Roeder G. S. Sex and the single cell: meiosis in yeast. Proc Natl Acad Sci U S A. 1995 Nov 7;92(23):10450–10456. doi: 10.1073/pnas.92.23.10450. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Scherthan H., Eils R., Trelles-Sticken E., Dietzel S., Cremer T., Walt H., Jauch A. Aspects of three-dimensional chromosome reorganization during the onset of human male meiotic prophase. J Cell Sci. 1998 Aug;111(Pt 16):2337–2351. doi: 10.1242/jcs.111.16.2337. [DOI] [PubMed] [Google Scholar]
  36. Scherthan H., Weich S., Schwegler H., Heyting C., Härle M., Cremer T. Centromere and telomere movements during early meiotic prophase of mouse and man are associated with the onset of chromosome pairing. J Cell Biol. 1996 Sep;134(5):1109–1125. doi: 10.1083/jcb.134.5.1109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Schwacha A., Kleckner N. Identification of double Holliday junctions as intermediates in meiotic recombination. Cell. 1995 Dec 1;83(5):783–791. doi: 10.1016/0092-8674(95)90191-4. [DOI] [PubMed] [Google Scholar]
  38. Shinohara A., Ogawa H., Matsuda Y., Ushio N., Ikeo K., Ogawa T. Cloning of human, mouse and fission yeast recombination genes homologous to RAD51 and recA. Nat Genet. 1993 Jul;4(3):239–243. doi: 10.1038/ng0793-239. [DOI] [PubMed] [Google Scholar]
  39. Shinohara A., Ogawa H., Ogawa T. Rad51 protein involved in repair and recombination in S. cerevisiae is a RecA-like protein. Cell. 1992 May 1;69(3):457–470. doi: 10.1016/0092-8674(92)90447-k. [DOI] [PubMed] [Google Scholar]
  40. Shinohara A., Ogawa T. Stimulation by Rad52 of yeast Rad51-mediated recombination. Nature. 1998 Jan 22;391(6665):404–407. doi: 10.1038/34943. [DOI] [PubMed] [Google Scholar]
  41. Smith K. N., Nicolas A. Recombination at work for meiosis. Curr Opin Genet Dev. 1998 Apr;8(2):200–211. doi: 10.1016/s0959-437x(98)80142-1. [DOI] [PubMed] [Google Scholar]
  42. Stassen N. Y., Logsdon J. M., Jr, Vora G. J., Offenberg H. H., Palmer J. D., Zolan M. E. Isolation and characterization of rad51 orthologs from Coprinus cinereus and Lycopersicon esculentum, and phylogenetic analysis of eukaryotic recA homologs. Curr Genet. 1997 Feb;31(2):144–157. doi: 10.1007/s002940050189. [DOI] [PubMed] [Google Scholar]
  43. Sung P. Function of yeast Rad52 protein as a mediator between replication protein A and the Rad51 recombinase. J Biol Chem. 1997 Nov 7;272(45):28194–28197. doi: 10.1074/jbc.272.45.28194. [DOI] [PubMed] [Google Scholar]
  44. Sung P., Robberson D. L. DNA strand exchange mediated by a RAD51-ssDNA nucleoprotein filament with polarity opposite to that of RecA. Cell. 1995 Aug 11;82(3):453–461. doi: 10.1016/0092-8674(95)90434-4. [DOI] [PubMed] [Google Scholar]
  45. Terasawa M., Shinohara A., Hotta Y., Ogawa H., Ogawa T. Localization of RecA-like recombination proteins on chromosomes of the lily at various meiotic stages. Genes Dev. 1995 Apr 15;9(8):925–934. doi: 10.1101/gad.9.8.925. [DOI] [PubMed] [Google Scholar]
  46. Weiner B. M., Kleckner N. Chromosome pairing via multiple interstitial interactions before and during meiosis in yeast. Cell. 1994 Jul 1;77(7):977–991. doi: 10.1016/0092-8674(94)90438-3. [DOI] [PubMed] [Google Scholar]
  47. Yoshida K., Kondoh G., Matsuda Y., Habu T., Nishimune Y., Morita T. The mouse RecA-like gene Dmc1 is required for homologous chromosome synapsis during meiosis. Mol Cell. 1998 Apr;1(5):707–718. doi: 10.1016/s1097-2765(00)80070-2. [DOI] [PubMed] [Google Scholar]

Articles from The Plant Cell are provided here courtesy of Oxford University Press

RESOURCES