Characterization of protein phosphatases in mouse oocytes

Okadaic acid (OA) enhances the resumption of meiosis in mouse oocytes, indicating that serine/threonine protein phosphatase-1 (PP1) and/or PP2A is involved. However, specific identification of PP1 and/or PP2A in mouse oocytes has not been reported. Here we demonstrate that fully grown germinal vesicle-intact (GVI) mouse oocytes contain mRNA corresponding to two isotypes of PP1, PP1alpha and PP1gamma. In addition, the transcript for PP2A was also present. At the protein level only PP1alpha and PP2A were recognized in fully grown GVI oocytes by Western blot analysis. Neither of the PP1gamma spliced variant proteins, PP1gamma1 and PP1gamma2, was detectable. Immunohistochemical analysis of ovarian tissue from gonadotropin-stimulated adult mice resulted in subcellular localization of both PP1alpha and PP2A, but not PP1gamma, in oocytes from all stages of folliculogenesis. In primordial oocytes, PP1alpha and PP2A were present in the cytoplasm. In more advanced stages of oogenesis, PP1alpha, although still present in the cytoplasm, was highly concentrated in the nucleus, whereas PP2A was predominantly cytoplasmic with a distinct reduction in the nuclear area. Both PP1alpha and PP2A were immunodetectable in oocytes during the prepubertal period. Eleven-day-old mouse oocytes, considered OA-insensitive and germinal vesicle breakdown (GVB)-incompetent, displayed both PP1alpha and PP2A predominantly in the cytoplasm. By 15 days of age mouse oocytes, which are beginning to acquire OA sensitivity and GVB competence, showed a relocation of PP1alpha into the nucleoplasm while PP2A remained predominantly cytoplasmic. This is the first specific identification of PP1alpha and PP2A in mouse oocytes. The differential localization of PP1alpha and PP2A, in addition to the relocation of PP1alpha during the acquisition of meiotic competence, suggests that these PPs have distinct regulatory roles during the resumption of meiosis.