Meiotic HORMA Domain Proteins Prevent Untimely Centriole
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Meiotic HORMA domain proteins prevent untimely PNAS PLUS centriole disengagement during Caenorhabditis elegans spermatocyte meiosis Mara Schvarzsteina,1, Divya Pattabiramana, Joshua N. Bembenekb, and Anne M. Villeneuvea,1 aDepartments of Developmental Biology and Genetics, Stanford University School of Medicine, Stanford, CA 94305; and bDepartment of Biochemistry, Cellular, and Molecular Biology, University of Tennessee, Knoxville, TN 37916 AUTHOR SUMMARY Sexual reproduction relies on protein family sharing a feature the production of complemen- Metaphase I wild type horma mutants called the HORMA domain, is a tary gametes that, together, crucial part of this strategy in contribute all the components C. elegans (2). HTP-1/2 localizes necessary for normal embryonic Anaphase I to the chromosome domains development. In addition to each where cohesion is retained gamete contributing a single during meiosis I and prevents haploid set of chromosomes, the Metaphase II the untimely separation of sister sperm and the egg (or oocyte) chromatids by locally inhibiting in many animal species also the removal of cohesin complexes fi provide the zygote, or newly or containing the meiosis-speci c fertilized single-cell embryo, Sperm subunit REC-8 by a cysteine with complementary compo- protease called separase. nents of the centrosome. Cen- The fact that meiosis involves trosomes are organelles that Zygote two rounds of cell division fol- male nucleate and help organize arrays pronucleus lowing a single round of DNA of microtubules within the cell. replication also necessitates a female fi Each centrosome contains either pronucleus modi cation of the centriole one or two cylindrical microtu- duplication cycle: During male bule structures called centrioles, Centriole pairs: engaged, disengaged DNA Centriole Microtubules meiosis in C. elegans,asin which recruit additional proteins mammals, centrioles normally CELL BIOLOGY termed pericentriolar material Fig. P1. Meiotic HORMA proteins limit centriole number in C. elegans undergo two rounds of duplica- (PCM) to form a functional zygotes by inhibiting centriole disengagement during meiosis II in tion. In nematodes, this results centrosome. A commonly used spermatocytes. Arrowheads highlight centriole foci marked by either in each haploid sperm contain- strategy for ensuring that gam- SAS-4 or SPD-2 antibodies. Widths of image panels: metaphase II, ing a single orthogonally en- 6.2 μm; sperm, 3.1 μm; zygote male pronuclei, 8.4 μm. etes provide complementary gaged centriole pair that will contributions of centrosome later disengage and duplicate components to the zygote is to dispose of either the centrioles or in the zygote to form the centrosomes of the first mitotic division. the PCM selectively by the end of gametogenesis (1). In the Because disengagement can license centriole duplication, sper- nematode Caenorhabditis elegans, as in humans, oocytes dispose matocytes must somehow prevent centriole disengagement at of the centrioles, whereas spermatocytes discard the PCM by meiosis II to ensure that sperm inherit a single engaged centriole the end of spermatogenesis. At fertilization, the oocyte con- pair. Indeed, in the current work, we show that inhibition of tributes the PCM and the sperm contributes the centriole pair centriole disengagement during meiosis II is critical to guarantee that, together, form the first centrosome of the embryo. There- the correct number of centrioles and centrosomes in the fore, maintaining the correct organization of centrioles during zygote (Fig. P1). Thus, prevention of centriole disengagement male meiosis is key to ensuring formation of a normal bipolar at meiosis II represents another important difference between mitotic spindle in the zygote. mitosis and meiosis. During mitotic cell cycles, key events, such as DNA replica- In this work, we investigate how proper centriole organization tion, chromosome segregation, centriole duplication, and cell is maintained during C. elegans male meiosis. Our results reveal division, are tightly coordinated and take place only once per an unanticipated role for members of the HORMA domain cycle. Further, like sister chromatids, centrioles separate during protein family in regulation of centriole disengagement during or soon after anaphase, resulting in two daughter cells that meiosis. Specifically, we used high-resolution immunofluores- each contain both a full genetic complement of chromosomes and a disengaged pair of centrioles. During meiosis, however, a single round of DNA replication is followed by two rounds Author contributions: M.S. and A.M.V. designed research; M.S. and D.P. performed of chromosome segregation: meiosis I, in which homologous research; M.S. and J.N.B. contributed new reagents/analytic tools; M.S. and A.M.V. chromosomes segregate away from each other but sisters stay analyzed data; and M.S. and A.M.V. wrote the paper. together, and meiosis II, in which sister chromatids segregate. The authors declare no conflict of interest. This meiotic pattern of chromosome segregation requires that This article is a PNAS Direct Submission. sister chromatid cohesion be released in two steps during the 1To whom correspondence may be addressed. E-mail: [email protected] or annev@ meiotic program, and it represents a major difference between stanford.edu. mitosis and meiosis. In previous work, we discovered that the See full research article on page E898 of www.pnas.org. HTP-1/2 protein, which is a member of a meiosis-specific Cite this Author Summary as: PNAS 10.1073/pnas.1213888110. www.pnas.org/cgi/doi/10.1073/pnas.1213888110 PNAS | March 5, 2013 | vol. 110 | no. 10 | 3727–3728 Downloaded by guest on September 27, 2021 cence imaging of WT and mutant C. elegans spermatocytes, ultimately destined for regulated separation. For mitotic cells, spermatids, and zygotes to identify a previously unrecognized it was previously suggested that the use of cohesin and separase role for C. elegans HORMA domain proteins HTP-1/2 and to regulate both sister chromatid cohesion and centriole en- HIM-3 in preventing centriole disengagement during the meiosis gagement might have evolved as a means to couple the two II division in spermatocytes. Our analysis revealed untimely types of separation events temporally during the cell cycle (3). centriole disengagement in him-3 and htp-1 htp-2 mutant sper- However, HORMA-dependent mechanisms operate to maintain matocytes, resulting in separated centrioles in sperm (Fig. P1). connections between sister chromatids during meiosis I (2) Further, an extra pair of centrosomes was detected in a subset and to maintain connections between centrioles during meiosis II of zygotes, suggesting that premature centriole disengagement (our study). In both cases, these mechanisms prevent inappro- may have enabled a single additional round of duplication priate separation events that could potentially impair the sub- (Fig. P1). We also provide evidence that these HORMA proteins sequent cell division. Thus, it is clear that parallel regulatory likely function to maintain centriole engagement in meiosis II logic, rather than temporal coordination, is the relevant under- fi by inhibiting separase-dependent removal of meiosis-speci c lying commonality that drove the dual use of this HORMA- cohesion complexes containing REC-8. Thus, our data suggest dependent cohesin maintenance strategy in regulating both that the same specialized meiotic mechanism that functions to chromosome and centriole separation during meiosis. prevent premature release of sister chromatid cohesion during meiosis I in C. elegans also functions to inhibit centriole sepa- 1. Manandhar G, Schatten H, Sutovsky P (2005) Centrosome reduction during gametogenesis ration at meiosis II, thereby ensuring that the zygote inherits the and its significance. Biol Reprod 72(1):2–13. appropriate complement of both chromosomes and centrioles. 2. Martinez-Perez E, et al. (2008) Crossovers trigger a remodeling of meiotic chromosome Our findings emphasize the fact that entire biological axis composition that is linked to two-step loss of sister chromatid cohesion. Genes Dev 22(20):2886–2901. subroutines can be recruited to accomplish distinct tasks that 3. Schöckel L, Möckel M, Mayer B, Boos D, Stemmann O (2011) Cleavage of cohesin require similar regulatory logic. In this case, the task is to rings coordinates the separation of centrioles and chromatids. Nat Cell Biol 13(8): maintain connections temporarily between structures that are 966–972. 3728 | www.pnas.org/cgi/doi/10.1073/pnas.1213888110 Schvarzstein et al. Downloaded by guest on September 27, 2021.