Pituitary gonadotropins perform essential functions in mammalian reproduction by stimulating gametogenesis and steroidogenesis in the ovaries and testicles. EZH2 is a histone methyltransferase that inhibits proliferation and aggravates apoptosis in stem cells subjected to pathological stimuli. However, the expression and molecular mechanisms of EZH2 in pituitary cells in vitro have not been extensively studied. In this study, the relative abundances of EZH2 mRNA (p< 0.01) and protein (p< 0.05) expression were larger in the pituitary cells of Hu sheep with relatively greater fecundity (GF) compared to those with lesser fecundity (LF). Loss-of-function examinations demonstrated thatEZH2gene knockdown led to an earlier induction of apoptosis in sheep pituitary cells (PCs). The relative abundance ofCASP3,CASP9, andBAXwas increased (p< 0.01), whileBCL2’s abundance was less decreased (p< 0.01) in PCs where there wasEZH2gene knockdown. Additionally, cell proliferation (p< 0.01) and viability (p< 0.01) were decreased in EZH2-knockdown sheep PCs, and the cell cycle was blocked compared to a negative control (NC). Notably,EZH2gene knockdown led to reduced abundances of gonadotropin subunit gene transcripts (FSHβ,p< 0.05) and reduced FSH release (p< 0.01) from PCs.EZH2gene knockdown led to reduced phosphorylation of AKT, ERK, and mTOR (p< 0.01). The results suggest that EZH2 regulates pituitary cell proliferation, apoptosis, and FSH secretion through modulation of the AKT/ERK signaling pathway, providing a foundation for further study of pituitary cell functions.