Nitric oxide regulates spindle dynamics to modulate the maturation of goat oocytes

Oocyte maturation is a complex and tightly regulated process. Nitric oxide (NO) has been implicated in the regulation of oocyte maturation, but its precise mechanism of action remains unclear. In this study, the inhibition of NO synthase activity using L-NMMA was found to impair meiosis and inhibit the maturation of goat oocytes. Further analysis revealed that the meiotic arrest induced by L-NMMA was primarily due to disruptions in metaphase I (MI) spindle dynamics. Specifically, L-NMMA treatment led to disorganized MI spindle assembly, abnormal chromosome alignment, elevated levels of microtubule acetylation, and sustained activation of the spindle assembly checkpoint. These defects were partially rescued by supplementation with the NO donor sodium nitroprusside (SNP). Proteomic analysis of oocytes from the CON and L-NMMA-treated groups identified a significant downregulation of KIF15 in the L-NMMA group. KIF15 has previously been established as a key regulator of spindle dynamics during oocyte maturation. Moreover, we demonstrated that the RhoA–ROCK signaling pathway modulated KIF15 protein expression in goat oocytes. Our results further showed that reduced NO levels increased RhoA phosphorylation, which in turn suppressed KIF15 expression and disrupted MI spindle dynamics. In conclusion, this study provides new insights into the role of NO in goat oocyte maturation, revealing that it regulates spindle dynamics through the RhoA–ROCK-KIF15 signaling axis.