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Abstract: The inhibitor Y-27632 is a specific selective inhibitor of Rho-associated protein kinases (ROCKs), which are downstream effectors of Rho guanosine triphosphatease (GTPases) and regulate Rho-associated cellular functions, including actin cytoskeletal organization. Little is known regarding the effects of Y-27632 on mammalian oocyte maturation. In the present study, we investigated the effects of Y-27632 on porcine oocyte meiosis and possible regulatory mechanisms of ROCK during porcine oocyte maturation. We found that ROCK accumulated not only at spindles, but also at the cortex in porcine oocytes. Y-27632 treatment reduced ROCK expression, and inhibited porcine oocyte meiotic maturation, which might be because of the impairment of actin expression and actin-related spindle positioning. Y-27632 treatment also disrupted the formation of actin cap and cortical granule-free domain, which further confirmed a spindle positioning failure. Thus, Y-27632 has significant effects on the meiotic competence of mammalian oocytes by reducing ROCK expression, and the regulation is related to its effects on actin-mediated spindle positioning. [Copyright &y& Elsevier]

PLK4, a polo-like kinase (PLK) family member that accumulates in the cytoplasm, has been identified as a crucial regulator of centriole formation. PLK4 also controls several essential cellular functions, including cytokinesis and gene expression. In this study, we investigated the expression and function of PLK4 during bovine oocyte meiotic maturation and subsequent embryo development. The PLK4 mRNA was detected in bovine oocytes at all developmental stages during meiotic maturation. Immunofluorescence staining showed that PLK4 protein exhibited a dynamic localization pattern in the oocyte cytoplasm during meiotic maturation, and fluorescence immunostaining markedly increased in metaphase II. When an interfering double-stranded RNA targeting PLK4 was injected into germinal vesicle–stage oocytes, PLK4 transcript levels decreased significantly in metaphase II oocytes (P < 0.05). The PLK4 knockdown caused spindle defects and chromosome misalignment and considerably reduced the amount of cortical and cytoplasmic actin. PLK4 was localized in the cytoplasm of early embryos, and PLK4 knockdown in germinal vesicle–stage oocytes led to failure in the early development of in vitro fertilized embryos (P < 0.05). Taken together, these results indicated that PLK4 plays crucial roles in bovine oocyte meiotic maturation and subsequent early embryo development. [ABSTRACT FROM AUTHOR]