Rap1b is a critical regulator of germinal vesicle breakdown in porcine oocytes

Germinal vesicle breakdown (GVBD) is a pivotal event in oocyte nuclear maturation, yet its underlying molecular mechanisms remain incompletely elucidated. This study showed that GVBD initiation in porcine oocytes occurred at 19 h of in vitro maturation, and this process was characterized by a significant increase in lamin A/C phosphorylation, prominent spindle assembly, and marked phosphorylation coupled with nuclear translocation of p38α MAPK (P < 0.05). Inhibition of p38 MAPK with SB203580 substantially suppressed p-p38α MAPK level, resulting in a significant decrease in GVBD rate, accompanied by notably diminished p-lamin A/C and impaired spindle assembly (P < 0.05). Single-cell transcriptome sequencing identified Rap1b, a small GTPase, as a potential upstream regulator of p38 MAPK pathway during GVBD. Further study demonstrated that Rap1b overexpression significantly enhanced both p-p38α MAPK and p-lamin A/C, promoted spindle assembly, and increased GVBD rate. Conversely, Rap1b knockdown markedly suppressed p-p38α MAPK and p-lamin A/C, disrupted spindle formation and decreased GVBD rate (P < 0.05). Further investigation revealed that Rap1b acted by promoting p-MKK6 to activate p38α MAPK signaling pathway. Collectively, these findings demonstrate that Rap1b is an essential regulator of oocyte GVBD by activating p38α MAPK signaling pathway. This work provides novel insights into the molecular regulation of oocyte meiotic resumption and offers a strategic basis for enhancing oocyte maturation.