Study on the Function of ID2 Gene in Granulosa Cells of Ovaries of Hetian Sheep and Its Correlation Analysis with Lambing Traits.

Reproductive efficiency is a key determinant of sheep productivity, yet Hetian sheep remain limited by relatively low fecundity despite their adaptability to harsh environments. The inhibitor of DNA binding 2 (ID2) gene is known to regulate cell proliferation and differentiation, but its specific role in sheep reproduction is not well understood. This study aimed to characterize the ID2 gene in Hetian sheep and to assess its functional association with ovarian granulosa cells and litter size. The coding sequence of ovine ID2 was cloned and analyzed using bioinformatics tools. Tissue-specific expression patterns were measured by quantitative PCR at different pubertal stages. A total of 157 ewes were genotyped to identify single-nucleotide polymorphisms (SNPs) and their association with litter size. Functional studies were performed by lentiviral overexpression of ID2 in granulosa cells, with effects evaluated using CCK-8 proliferation assays, ELISA for hormone secretion, and RT-qPCR for related gene expression. ID2 was highly expressed in the ovary, particularly during puberty. Four SNPs (g.18202368 A>T, g.18202372 G>A, g.18202431 G>C, g.18202472 G>C) were significantly associated with increased litter size. Overexpression of ID2 promoted granulosa cell proliferation, increased progesterone, decreased estradiol, and altered expression of key genes in the TGF-β/BMP-SMAD signaling pathway. The ID2 gene plays a crucial role in ovarian function and reproductive regulation in Hetian sheep. Its polymorphisms and functional impact on granulosa cells suggest that ID2 is a promising candidate gene for marker-assisted selection to improve reproductive efficiency in sheep.