Granulosa cell-layer stiffening prevents the granulosa cells from escaping the post-ovulatory follicle

Abstract

Ovulation is necessary for successful reproduction. After ovulation, cumulus cells and oocytes are released, while granulosa cells (GCs) remain trapped within the post-ovulatory follicle to form the corpus luteum. However, the mechanism underlying GC confinement has long been unclear. Here, we providein vitroandin vivoevidence demonstrating that the stiffening of GC-layer as an evolutionarily conserved mechanism that hinders GCs from escaping the post-ovulatory follicles. Spatial transcriptome analysis reveals that the assembly of focal adhesions is primarily responsible for this stiffening. Disrupting focal adhesion assembly through RNA interference results in the release of GCs from the post-ovulatory follicle, leading to the formation of an aberrant corpus luteum with reduced cell density and cavities. We also uncover that theLH (hCG) -cAMP-PKA-CREBsignaling axis stimulates focal adhesion assembly and induce GC-layer stiffening. Our findings introduce a novel concept of “GC-layer stiffening”, which offers valuable insights into the factors that prevent GCs escape from the post-ovulatory follicle.