PRSS23 Promotes Ovarian Follicular Atresia in Wuding Chickens by Coordinately Suppressing Steroidogenesis and PI3K/AKT/mTOR Survival Signaling

<b>Background</b>: Broodiness is a major limiting factor for reproductive efficiency in indigenous avian breeds, a phenomenon underpinned physiologically by granulosa cell (GC) apoptosis and subsequent follicular atresia. While Serine Protease 23 (PRSS23) has been implicated in mammalian ovarian remodeling, its specific regulatory function in avian follicular dynamics remains elusive. <b>Methods</b>: Utilizing the Wuding chicken-an indigenous breed distinguished by robust environmental adaptability but compromised by high broodiness frequency-as a biological model, this study dissected the molecular mechanism of PRSS23-mediated follicular regression. We cloned the complete coding sequence of the Wuding chicken <i>PRSS23</i> gene, characterized its spatiotemporal expression profile, and interrogated its function in primary GCs via gain- and loss-of-function assays. <b>Results</b>: RT-qPCR analysis revealed that <i>PRSS23</i> is differentially expressed across the hypothalamic-pituitary-ovarian (HPO) axis, with ovarian expression being significantly upregulated during the broody period compared to the laying period. Mechanistically, <i>PRSS23</i> overexpression significantly downregulated the expression of follicle-stimulating hormone receptor (<i>FSHR</i>) and key steroidogenic enzymes (<i>STAR</i>, <i>CYP19A1</i>, <i>HSD3β1</i>), thereby suppressing the expression of genes governing the biosynthesis potential of progesterone and estradiol. Concurrently, <i>PRSS23</i> overexpression was associated with transcriptional repression of components of the PI3K/AKT/mTOR signaling cascade; this transcriptional regulation further induced cell cycle arrest at the G0/G1 phase, and activated the mitochondrial apoptotic pathway characterized by <i>BAX</i> upregulation and <i>BCL2</i> downregulation. Conversely, siRNA-mediated knockdown of <i>PRSS23</i> alleviated these inhibitory effects, promoting GC proliferation and survival. <b>Conclusions</b>: These findings establish <i>PRSS23</i> as a pivotal pro-atretic factor in Wuding chickens, driving ovarian atrophy through the dual transcriptional-level inhibition of steroidogenesis and survival signaling pathways. This study identifies a potential molecular target for marker-assisted selection programs aimed at attenuating broodiness while preserving the superior meat quality traits of indigenous poultry.