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The Zebrafish Meiotic Cohesin Complex Protein Smc1b Is Required for Key Events in Meiotic Prophase I.

ABSTRACT: The eukaryotic structural maintenance of chromosomes (SMC) proteins are involved in key processes of chromosome structure and dynamics. SMC1β was identified as a component of the meiotic cohesin complex in vertebrates, which aids in keeping sister chromatids together prior to segregation in meiosis II and is involved in association of homologous chromosomes in meiosis I. The role of SMC1β in meiosis has primarily been studied in mice, where mutant male and female mice are infertile due to germ cell arrest at pachytene and metaphase II stages, respectively. Here, we investigate the function of zebrafish Smc1b to understand the role of this protein more broadly in vertebrates. We found that zebrafish smc1b is necessary for fertility and has important roles in meiosis, yet has no other apparent roles in development. Therefore, smc1b functions primarily in meiosis in both fish and mammals. In zebrafish, we showed that smc1b mutant spermatocytes initiated telomere clustering in leptotene, but failed to complete this process and progress into zygotene. Furthermore, mutant spermatocytes displayed a complete failure of synapsis between homologous chromosomes and homolog pairing only occurred at chromosome ends. Interestingly, meiotic DNA double strand breaks occurred in the absence of Smc1b despite failed pairing and synapsis. Overall, our findings point to an essential role of Smc1b in the leptotene to zygotene transition during zebrafish spermatogenesis. In addition, ovarian follicles failed to form in smc1b mutants, suggesting an essential role in female meiosis as well. Our results indicate that there are some key differences in Smc1b requirement in meiosis among vertebrates: while Smc1b is not required for homolog pairing and synapsis in mice, it is essential for these processes in zebrafish.

SUBMITTER: Islam KN

PROVIDER: S-EPMC8381726 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The cohesin complexes play a key role in chromosome segregation during both mitosis and meiosis. They establish sister chromatid cohesion between duplicating DNA molecules during S-phase, but they also have an important role during postreplicative double-strand break repair in mitosis, as well as during recombination between homologous chromosomes in meiosis. An additional function in meiosis is related to the sister kinetochore cohesion, so they can be pulled by microtubules to the same pole at anaphase I. Data about the dynamics of cohesin subunits during meiosis are scarce; therefore, it is of great interest to characterize how the formation of the cohesin complexes is achieved in order to understand the roles of the different subunits within them. We have investigated the spatio-temporal distribution of three different cohesin subunits in prophase I grasshopper spermatocytes. We found that structural maintenance of chromosome protein 3 (SMC3) appears as early as preleptotene, and its localization resembles the location of the unsynapsed axial elements, whereas radiation-sensitive mutant 21 (RAD21) (sister chromatid cohesion protein 1, SCC1) and stromal antigen protein 1 (SA1) (sister chromatid cohesion protein 3, SCC3) are not visualized until zygotene, since they are located in the synapsed regions of the bivalents. During pachytene, the distribution of the three cohesin subunits is very similar and all appear along the trajectories of the lateral elements of the autosomal synaptonemal complexes. However, whereas SMC3 also appears over the single and unsynapsed X chromosome, RAD21 and SA1 do not. We conclude that the loading of SMC3 and the non-SMC subunits, RAD21 and SA1, occurs in different steps throughout prophase I grasshopper meiosis. These results strongly suggest the participation of SMC3 in the initial cohesin axis formation as early as preleptotene, thus contributing to sister chromatid cohesion, with a later association of both RAD21 and SA1 subunits at zygotene to reinforce and stabilize the bivalent structure. Therefore, we speculate that more than one cohesin complex participates in the sister chromatid cohesion at prophase I.

Project description:Meiotic chromosome segregation relies on homologous chromosomes being linked by at least one crossover, the obligate crossover. Homolog pairing, synapsis and meiosis specific DNA repair mechanisms are required for crossovers but how they are coordinated to promote the obligate crossover is not well understood. PCH-2 is a highly conserved meiotic AAA+-ATPase that has been assigned a variety of functions; whether these functions reflect its conserved role has been difficult to determine. We show that PCH-2 restrains pairing, synapsis and recombination in C. elegans. Loss of pch-2 results in the acceleration of synapsis and homolog-dependent meiotic DNA repair, producing a subtle increase in meiotic defects, and suppresses pairing, synapsis and recombination defects in some mutant backgrounds. Some defects in pch-2 mutants can be suppressed by incubation at lower temperature and these defects increase in frequency in wildtype worms grown at higher temperature, suggesting that PCH-2 introduces a kinetic barrier to the formation of intermediates that support pairing, synapsis or crossover recombination. We hypothesize that this kinetic barrier contributes to quality control during meiotic prophase. Consistent with this possibility, defects in pch-2 mutants become more severe when another quality control mechanism, germline apoptosis, is abrogated or meiotic DNA repair is mildly disrupted. PCH-2 is expressed in germline nuclei immediately preceding the onset of stable homolog pairing and synapsis. Once chromosomes are synapsed, PCH-2 localizes to the SC and is removed in late pachytene, prior to SC disassembly, correlating with when homolog-dependent DNA repair mechanisms predominate in the germline. Indeed, loss of pch-2 results in premature loss of homolog access. Altogether, our data indicate that PCH-2 coordinates pairing, synapsis and recombination to promote crossover assurance. Specifically, we propose that the conserved function of PCH-2 is to destabilize pairing and/or recombination intermediates to slow their progression and ensure their fidelity during meiotic prophase.

Project description:The centromere is an epigenetic mark that is a loading site for the kinetochore during meiosis and mitosis. This mark is characterized by the H3 variant CENP-A, known as CID in Drosophila . In Drosophila , CENP-C is critical for maintaining CID at the centromeres and directly recruits outer kinetochore proteins after nuclear envelope break down. It is not known, however, if these two functions require the same CENP-C molecules. Furthermore, in Drosophila and many other metazoan oocytes, centromere maintenance and kinetochore assembly are separated by an extended prophase. Consistent with studies in mammals, CID is relatively stable and does not need to be expressed during prophase to remain at high levels in metaphase I of meiosis. Expression of CID during prophase can even be deleterious, causing ectopic localization to non-centromeric chromatin, abnormal meiosis and sterility. In contrast to CID, maintaining high levels of CENP-C requires expression during prophase. Confirming the importance of this loading, we found CENP-C prophase loading is required for multiple meiotic functions. In early meiotic prophase, CENP-C loading is required for sister centromere cohesion and centromere clustering. In late meiotic prophase, CENP-C loading is required to recruit kinetochore proteins. CENP-C is one of the few proteins identified in which expression during prophase is required for meiotic chromosome segregation. An implication of these results is that the failure to maintain recruitment of CENP-C during the extended prophase in oocytes would result in chromosome segregation errors in oocytes.Author summaryMeiosis in oocytes of diverse organisms, including humans and Drosophila, is characterized by a long prophase pause and a cell cycle arrest in meiosis I or meiosis II. These pauses could be a challenge for centromeres, whose components are replenished during G1, and then must remain with the chromosomes until the meiotic divisions. We have investigated the stability, prophase dynamics and function of centromere protein CENP-C. We show that CENP-C expression and loading onto centromeres during prophase is required for multiple meiotic functions. In contrast, the expression of other centromere partners CID/CENP-A and CAL1 is not required during meiotic prophase. Furthermore, expression of CID during prophase can be deleterious and result in ectopic kinetochore formation. CENP-C loading in prophase is required for sister centromere cohesion and kinetochore assembly. Our results provide the first description of CENP-C dynamics during meiosis and show that prophase expression is required for oocyte spindle assembly and function. CENP-C is among a small number of proteins that are required for the meiotic divisions but are loaded prior to prometaphase. Failure to maintain these proteins during the long prophase of oocyte meiosis may contribute to the increased aneuploidy associated with advanced maternal age.

Project description:the synaptonemal complex (SC) links two meiotic prophase chromosomal events: homolog pairing and crossover recombination. SC formation involves the multimeric assembly of coiled-coil proteins (Zip1 in budding yeast) at the interface of aligned homologous chromosomes. However, SC assembly is indifferent to homology and thus is normally regulated such that it occurs only subsequent to homology recognition. Assembled SC structurally interfaces with and influences the level and distribution of interhomolog crossover recombination events. Despite its involvement in dynamic chromosome behaviors such as homolog pairing and recombination, the extent to which SC, once installed, acts as an irreversible tether or maintains the capacity to remodel is not clear. Experiments presented here reveal insight into the dynamics of the full-length SC in budding yeast meiotic cells. We demonstrate that Zip1 continually incorporates into previously assembled synaptonemal complex during meiotic prophase. Moreover, post-synapsis Zip1 incorporation is sufficient to rescue the sporulation defect triggered by SCs built with a mutant version of Zip1, Zip1-4LA. Post-synapsis Zip1 incorporation occurs initially with a non-uniform spatial distribution, predominantly associated with Zip3, a component of the synapsis initiation complex that is presumed to mark a subset of crossover sites. A non-uniform dynamic architecture of the SC is observed independently of (i) synapsis initiation components, (ii) the Pch2 and Pph3 proteins that have been linked to Zip1 regulation, and (iii) the presence of a homolog. Finally, the rate of SC assembly and SC central region size increase in proportion to Zip1 copy number; this and other observations suggest that Zip1 does not exit the SC structure to the same extent that it enters. Our observations suggest that, after full-length assembly, SC central region exhibits little global turnover but maintains differential assembly dynamics at sites whose distribution is patterned by a recombination landscape.

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The Zebrafish Meiotic Cohesin Complex Protein Smc1b Is Required for Key Events in Meiotic Prophase I.

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