RESUMO
Chromobox homolog 2 (CBX2), a key member of the polycomb group (PcG) family, is essential for gonadal development in mammals. A functional deficiency or genetic mutation in cbx2 can lead to sex reversal in mice and humans. However, little is known about the function of cbx2 in gonadal development in fish. In this study, the cbx2 gene was identified in medaka, which is a model species for the study of gonadal development in fish. Transcription of cbx2 was abundant in the gonads, with testicular levels relatively higher than ovarian levels. In situ hybridization (ISH) revealed that cbx2 mRNA was predominately localized in spermatogonia and spermatocytes, and was also observed in oocytes at stages I, II, and III. Furthermore, cbx2 and vasa (a marker gene) were co-localized in germ cells by fluorescent in situ hybridization (FISH). After cbx2 knockdown in the gonads by RNA interference (RNAi), the sex-related genes, including sox9 and foxl2, were influenced. These results suggest that cbx2 not only plays a positive role in spermatogenesis and oogenesis but is also involved in gonadal differentiation through regulating the expression levels of sex-related genes in fish.
Assuntos
Proteínas de Peixes/genética , Gônadas/metabolismo , Oryzias/genética , Complexo Repressor Polycomb 1/genética , Sequência de Aminoácidos , Animais , Feminino , Proteínas de Peixes/antagonistas & inibidores , Proteínas de Peixes/classificação , Proteínas de Peixes/metabolismo , Proteína Forkhead Box L2/antagonistas & inibidores , Proteína Forkhead Box L2/genética , Proteína Forkhead Box L2/metabolismo , Gônadas/crescimento & desenvolvimento , Masculino , Oryzias/crescimento & desenvolvimento , Filogenia , Complexo Repressor Polycomb 1/antagonistas & inibidores , Complexo Repressor Polycomb 1/classificação , Complexo Repressor Polycomb 1/metabolismo , Interferência de RNA , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Fatores de Transcrição SOX9/antagonistas & inibidores , Fatores de Transcrição SOX9/genética , Fatores de Transcrição SOX9/metabolismo , Alinhamento de Sequência , Espermatócitos/metabolismo , Espermatogônias/metabolismoRESUMO
Polycomb repressive complex 1 (PRC1) is required for ubiquitination of histone H2A lysine 119, an epigenetic mark associated with repression of genes important in developmental regulation. The E3 ligase activity of PRC1 resides in the RING1A/B subunit when paired with one of six PCGF partners. The best known of these is the oncogene BMI1/PCGF4. We find that canonical PRC1 E3 ligases such as PCGF4-RING1B have intrinsically very low enzymatic activity compared with non-canonical PRC1 RING dimers. The structure of a high-activity variant in complex with E2 (PCGF5-RING1B-UbcH5c) reveals only subtle differences from an earlier PCGF4 complex structure. However, two charged residues present in the modelled interface with E2-conjugated ubiquitin prove critical: in BMI1/PCGF4, these residues form a salt bridge that may limit efficient ubiquitin transfer. The intrinsically low activity of the PCGF4-RING1B heterodimer is offset by a relatively favourable interaction with nucleosome substrates, resulting in an efficient site-specific monoubiquitination.