组蛋白H3Ser10磷酸化对猪卵母细胞减数分裂的影响

Effect of histone H3Ser10 phophorylation on oocyte meiotic in porcine

目的组蛋白磷酸化修饰在卵母细胞减数分裂过程中作用机制的研究较少。文中旨在探讨猪卵母细胞中组蛋白H3的磷酸化模式及对卵母细胞减数分裂过程的调控。方法首先采用免疫组化法鉴定组蛋白H3Ser10(H 3S10)磷酸化在猪卵母细胞减数分裂过程的表达情况,随后体外成熟猪卵母细胞,将卵丘卵母细胞复合体(COCs)分为4个组,一组正常培养作为对照组,另外3组分别添加5、10、30μmol/L ZM447439的成熟液,体外培养27 h,分别为5、10、30μmol/L ZM447439处理组。统计卵母细胞在减数分裂各个时期所占的比例,并进一步检测猪卵母细胞组蛋白H3S10磷酸化的表达模式及蛋白激酶Aurora B基因表达的情况。结果与GVBD期组蛋白H3S10磷酸化水平比较,MI期和AI期明显升高(P<0.05),AI期较MⅡ期明显升高(P<0.05)。与对照组相比,10、30μmol/L ZM447439处理组GVBD期卵母细胞比例[(32.14±0.51)%、(95.34±0.59)%]较对照组[(2.56±0.03)%]增加(P<0.05),MI[(66.88±0.13)%、(4.66±0.04)%]较对照组[(87.42±0.14)%]明显下降(P<0.05)、AI期[(1.01±0.03)%、(0.00±0.00)%]较对照组[(10.02±0.21)%]明显下降(P<0.05)。与对照组卵母细胞发生H3S10去磷酸化修饰比例(0)比较,10μmol/L、30μmol/L ZM447439处理组[(35.2±0.39)%、(95.4±0.65)%]明显升高(P<0.05)。与对照组相比,10、30μmol/L ZM447439处理组Aurora B基因的相对表达量显著降低(P<0.05)。结论组蛋白H3S10磷酸化修饰在哺乳动物卵母细胞成熟过程起着一定作用。Aurora B激酶抑制剂ZM447439处理引起组蛋白H3S10磷酸化水平及Aurora-B激酶的表达降低导致卵母细胞成熟障碍。

Objective The mechanism of histone phosphorylation modification in oocyte meiosis is less studied.This study is designed to investigate the pattern of histone H3 phophorylation and regulation of maturation process in the porcine oocytes.Methods The histone H3Ser10(H3S10)phosphorylation expression was examined on the porcine oocyte meiotic process.The porcine cumulus oocyte complexes(COCs)were divided into four groups,one group was cultured as control group,and the other 3 groups were supplemented with 5,10,and 30μmol/L ZM447439,and cultured in vitro for 27 h,respectively,5,10,and 30μmol/L ZM4474349 treatment group.The proportion of each meiotic stage was counted.The phosphorylation pattern of histone H3S10 and the expression level of protein kinase Aurora B were detected at the porcine oocytes.Results Compared with histone H3S10 phosphorylation level of oocyte GVBD phase,the MI and AI phases were significantly increased(P<0.05),and H3S10 phosphorylation level of AI phase was remarkedly higher than that of MII phase(P<0.05).Compared with the control group,the proportion of oocytes at the GVBD phase in the 10 and 30μmol/L ZM4447439 treatment group[(32.14±0.51)%,(95.34±0.59)%]was higher than that of the control group[(2.56±0.03)%,P<0.05],the proportion of oocytes at the MI phase[(66.88±0.13)%,(4.66±0.04)%]significantly decreased than that of the control group[(87.42±0.14)%,P<0.05],and the proportion of oocytes at the AI stage[(1.01±0.03)%,(0.000±0.00)%]significantly decreased compared with the control group[(10.02±0.21)%,P<0.05].Compared with the control group(0),oocytes H3S10 dephosphorylation modification ratio in the 10μmol/L and 30μmol/L ZM4474349 treatment group[(35.2±0.39)%,(95.4±0.65)%]significantly increased(P<0.05).Compared with the control group,the relative expression level of Aurora B in the 10 and 30μmol/L ZM4447439 treatment group was significantly reduced(P<0.05).Conclusion Hstone H3S10 phosphorylation plays arolein the maturation of mammalian oocytes.AuroraB kinase inhibitors(ZM447439)treatment can reduce H3S10 phosphorylation and Aurora B expression level and lead to oocytesmaturation disorder.