RESUMO
During meiosis, paired homologous chromosomes (homologs) become linked via the synaptonemal complex (SC) and crossovers. Crossovers mediate homolog segregation and arise from self-inflicted double-strand breaks (DSBs). Here, we identified a role for the proteasome, the multisubunit protease that degrades proteins in the nucleus and cytoplasm, in homolog juxtaposition and crossing over. Without proteasome function, homologs failed to pair and instead remained associated with nonhomologous chromosomes. Although dispensable for noncrossover formation, a functional proteasome was required for a coordinated transition that entails SC assembly between longitudinally organized chromosome axes and stable strand exchange of crossover-designated DSBs. Notably, proteolytic core and regulatory proteasome particles were recruited to chromosomes by Zip3, the ortholog of mammalian E3 ligase RNF212, and SC protein Zip1 . We conclude that proteasome functions along meiotic chromosomes are evolutionarily conserved.
Assuntos
Troca Genética , Meiose/fisiologia , Proteínas Nucleares/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Complexo Sinaptonêmico/enzimologia , Ubiquitina-Proteína Ligases/metabolismo , Núcleo Celular/enzimologia , Pareamento Cromossômico , Cromossomos Fúngicos/metabolismo , Inibidores de Cisteína Proteinase/farmacologia , Citoplasma/enzimologia , Quebras de DNA de Cadeia Dupla , Evolução Molecular , Leupeptinas/farmacologia , Meiose/genética , Proteínas Nucleares/genética , Complexo de Endopeptidases do Proteassoma/genética , Proteólise , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/fisiologia , Proteínas de Saccharomyces cerevisiae/genética , Complexo Sinaptonêmico/genética , Ubiquitina-Proteína Ligases/genéticaRESUMO
The Japanese flounder (Paralichthys olivaceus) is a teleost fish with an XX/XY sex determination system. XX flounder can be induced to develop into phenotypic females or males, by rearing them at 18°C or 27°C, respectively, during the sex differentiation period. Therefore, the flounder provides an excellent model to study the molecular mechanisms underlying temperature-dependent sex determination. We previously showed that cortisol, the major glucocorticoid produced by the interrenal cells in teleosts, causes female-to-male sex reversal by directly suppressing mRNA expression of ovary-type aromatase (cyp19a1), a steroidogenic enzyme responsible for the conversion of androgens to estrogens in the gonads. Furthermore, an inhibitor of cortisol synthesis prevented masculinization of XX flounder at 27°C, suggesting that masculinization by high temperature is due to the suppression of cyp19a1 mRNA expression by elevated cortisol levels during gonadal sex differentiation in the flounder. In the present study, we found that exposure to high temperature during gonadal sex differentiation upregulates the mRNA expression of retinoid-degrading enzyme (cyp26b1) concomitantly with masculinization of XX gonads and delays meiotic initiation of germ cells. We also found that cortisol induces cyp26b1 mRNA expression and suppresses specific meiotic marker synaptonemal complex protein 3 (sycp3) mRNA expression in gonads during the sexual differentiation. In conclusion, these results suggest that exposure to high temperature induces cyp26b1 mRNA expression and delays meiotic initiation of germ cells by elevating cortisol levels during gonadal sex differentiation in Japanese flounder.
Assuntos
Sistema Enzimático do Citocromo P-450/biossíntese , Linguado/crescimento & desenvolvimento , Células Germinativas/crescimento & desenvolvimento , Temperatura Alta , Meiose , Caracteres Sexuais , Diferenciação Sexual , Animais , Clonagem Molecular , Sistema Enzimático do Citocromo P-450/classificação , Sistema Enzimático do Citocromo P-450/genética , Feminino , Linguado/genética , Linguado/metabolismo , Células Germinativas/efeitos dos fármacos , Células Germinativas/enzimologia , Hidrocortisona/farmacologia , Masculino , Filogenia , RNA Mensageiro/biossíntese , Ácido Retinoico 4 Hidroxilase , Complexo Sinaptonêmico/enzimologiaRESUMO
The proper execution of premeiotic S phase is essential to both the maintenance of genomic integrity and accurate chromosome segregation during the meiotic divisions. However, the regulation of premeiotic S phase remains poorly defined in metazoa. Here, we identify the p21(Cip1)/p27(Kip1)/p57(Kip2)-like cyclin-dependent kinase inhibitor (CKI) Dacapo (Dap) as a key regulator of premeiotic S phase and genomic stability during Drosophila oogenesis. In dap(-/-) females, ovarian cysts enter the meiotic cycle with high levels of Cyclin E/cyclin-dependent kinase (Cdk)2 activity and accumulate DNA damage during the premeiotic S phase. High Cyclin E/Cdk2 activity inhibits the accumulation of the replication-licensing factor Doubleparked/Cdt1 (Dup/Cdt1). Accordingly, we find that dap(-/-) ovarian cysts have low levels of Dup/Cdt1. Moreover, mutations in dup/cdt1 dominantly enhance the dap(-/-) DNA damage phenotype. Importantly, the DNA damage observed in dap(-/-) ovarian cysts is independent of the DNA double-strands breaks that initiate meiotic recombination. Together, our data suggest that the CKI Dap promotes the licensing of DNA replication origins for the premeiotic S phase by restricting Cdk activity in the early meiotic cycle. Finally, we report that dap(-/-) ovarian cysts frequently undergo an extramitotic division before meiotic entry, indicating that Dap influences the timing of the mitotic/meiotic transition.