RESUMO
Mammalian early embryos maintain accurate genome integrity for proper development within a programmed timeline despite constant assaults on their DNA by replication, DNA demethylation and genetic defects transmitted from germ cells. However, how genome integrity is safeguarded during mammalian early embryonic development remains unclear. BCAS2 (breast carcinoma amplified sequence 2), a core component of the PRP19 complex involved in pre-mRNA splicing, plays an important role in the DNA damage response through the RPA complex, a key regulator in the maintenance of genome integrity. Currently, the physiological role of BCAS2 in mammals is unknown. We now report that BCAS2 responds to endogenous and exogenous DNA damage in mouse zygotes. Maternal depletion of BCAS2 compromises the DNA damage response in early embryos, leading to developmental arrest at the two- to four-cell stage accompanied by the accumulation of damaged DNA and micronuclei. Furthermore, BCAS2 mutants that are unable to bind RPA1 fail in DNA repair during the zygotic stage. In addition, phosphorylated RPA2 cannot localise to the DNA damage sites in mouse zygotes with disrupted maternal BCAS2. These data suggest that BCAS2 might function through the RPA complex during DNA repair in zygotes. Together, our results reveal that maternal BCAS2 maintains the genome integrity of early embryos and is essential for female mouse fertility.
Assuntos
Reparo do DNA/fisiologia , Desenvolvimento Embrionário/fisiologia , Fertilidade/fisiologia , Instabilidade Genômica/fisiologia , Complexos Multiproteicos/metabolismo , Proteínas de Neoplasias/metabolismo , Animais , Western Blotting , Reparo do DNA/genética , Desenvolvimento Embrionário/genética , Feminino , Fertilidade/genética , Marcação de Genes , Instabilidade Genômica/genética , Marcação In Situ das Extremidades Cortadas , Camundongos , Microscopia de Fluorescência , Proteínas Associadas à Matriz Nuclear/metabolismo , Gravidez , Fatores de Processamento de RNA , Reação em Cadeia da Polimerase em Tempo Real , Proteína de Replicação A/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Maternal effect genes play critical roles in early embryogenesis of model organisms where they have been intensively investigated. However, their molecular function in mammals remains largely unknown. Recently, we identified a subcortical maternal complex (SCMC) that contains four proteins encoded by maternal effect genes (Mater, Filia, Floped and Tle6). Here we report that TLE6, similar to FLOPED and MATER, stabilizes the SCMC and is necessary for cleavage beyond the two-cell stage of development. We document that the SCMC is required for formation of the cytoplasmic F-actin meshwork that controls the central position of the spindle and ensures symmetric division of mouse zygotes. We further demonstrate that the SCMC controls formation of the actin cytoskeleton specifically via Cofilin, a key regulator of F-actin assembly. Our results provide molecular insight into the physiological function of TLE6, its interaction with the SCMC and their roles in the symmetric division of the zygote in early mouse development.
Assuntos
Actinas/metabolismo , Antígenos/metabolismo , Divisão Celular/fisiologia , Proteínas do Ovo/metabolismo , Oócitos/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas Repressoras/genética , Zigoto/metabolismo , Citoesqueleto de Actina/metabolismo , Fatores de Despolimerização de Actina/metabolismo , Animais , Proteínas Correpressoras , Desenvolvimento Embrionário , Camundongos , Camundongos Knockout , Mitose , Proteínas/metabolismo , Proteínas Repressoras/metabolismoRESUMO
OBJECTIVE: To investigate the differential expression and regulation of Gstm2 in mouse uterus during early pregnancy. DESIGN: Experimental animal study. SETTING: University research laboratory. ANIMAL(S): Sexually mature female Kunming white strain mice. INTERVENTION(S): Delayed and activated implantation, pseudopregnancy, hormonal treatment. MAIN OUTCOME MEASURE(S): The expression of Gstm2 mRNA was detected by in situ hybridization and reverse-transcription polymerase chain reaction (RT-PCR). RESULT(S): By in situ hybridization, there were a low level of Gstm2 expression in luminal epithelium on day 3 and a strong level in the luminal epithelium on day 4 during early pregnancy. The expression pattern of Gstm2 in the pseudopregnant uterus was similar to that during early pregnancy. By RT-PCR, Gstm2 was strongly detected in the uteri on days 3 and 4 of pregnancy and pseudopregnancy. Gstm2 expression was strongly detected in the luminal epithelium under delayed implantation, but not seen after delayed implantation was activated by estrogen. In the ovariectomized mouse uterus, Gstm2 expression was strongly up-regulated by progesterone via progesterone receptor. CONCLUSION(S): The results showed that Gstm2 was highly expressed in the uterine luminal epithelium during preimplantation period and up-regulated by progesterone.
