MMP2基因启动子低甲基化延缓母猪初情启动的机制研究

初情启动是后备母猪性成熟和获得繁殖能力的标志，但研究表明，约14%的后备母猪因初情启动延缓而被淘汰，影响猪场的经济效益。项目申请人前期发现，MMP2启动子区低DNA甲基化状态可能影响E2/ERα对MMP2基因的转录调控，进而诱导颗粒细胞过度凋亡和有腔卵泡过量闭锁，最终导致母猪初情启动延缓的发生。本项目拟在前期基础上，以初情启动正常和延缓母猪的卵巢为实验材料，利用组织切片和bisulfite sequencing PCR等方法，明确MMP2基因与颗粒细胞凋亡、卵泡闭锁及初情启动延缓之间的关系；然后，在细胞和组织水平，采用过表达、siRNA、ChIP和启动子定点突变等方法，研究MMP2启动子区低DNA甲基化影响E2/ERα调控MMP2的转录表达，及其影响颗粒细胞功能和卵泡生长的作用机制。研究结果将从理论上揭示MMP2基因延缓母猪初情启动的作用机制，并为选育性成熟早的母猪提供理论依据。

In gilts, the initiation of puberty indicates the sexual maturation and reproductive competence. However, previous studies suggest that about 14% of gilts are culled before being served due to the delayed puberty, which is consequent in a heavily economic burden on the modern commercial farms. Our previous results revealed that the DNA methylation of Matrix metalloproteinases 2 (MMP2) promoter could block the transcriptional regulation of E2/ERα on MMP2, induce the excessive apoptosis of granulosa cells and follicular atresia, then delay the onset of puberty in gilts. Therefore, in this project, we first try to identify the relationship among MMP2, granulosa cell apoptosis, follicular atresia, and delayed puberty by using the ovaries from the normal and delayed gilts. Then we explore and determine the mechanism that how the DNA methylation of MMP2 promoter affect the transcriptional regulation of E2/ERα on MMP2, using the porcine ovarian granulosa cells and cultured follicles. We further investigate the influences of this mechanism on the proliferation and apoptosis of granulosa cells and follicular growth. These works could not only improve the theoretical resource available for better understanding the mechanism for the emergence of DP, but also provide valuable insights for the selection of early sexual maturation in gilts.