猪早期孤雌胚组蛋白乙酰化与组蛋白去乙酰化酶基因mRNA表达水平相关性研究

摘要

[目的]探讨猪孤雌胚早期发育过程中组蛋白H4K5乙酰化水平与组蛋白去乙酰化酶(HDAC1)基因mRNA 表达水平之间的关系.[方法]收集早期猪孤雌胚(2-细胞,4-细胞、桑椹胚和囊胚),运用细胞免疫组化和激光共聚焦显微镜,通过检测这些胚胎的相对荧光强度来评判其H4K5的乙酰化水平;运用实时定量PCR的方法检测猪早期孤雌胚发育过程中HDAC1基因mRNA表达的动态变化.[结果]从2-细胞到囊胚开始,其H4K5的乙酰化均在细胞核内表达,且各阶段表达量逐步提高,至囊胚期达最高(分别为1124.77土127.78,1305.81土184.23,1795.19-±318.67及2149.63土529.47),差异显著(P<0.05);从2-细胞到囊胚,其HDAC1基因mRNA表达逐步降低(分别为1.00土0、0.91土0.009、0.85土0.008及0.67土0.006),各阶段差异显著(P<0.05).[结论]从2-细胞到囊胚,其H4K5的乙酰化水平与HDAC1基因mRNA表达水平之间呈负相关.%[Objective]The aims of this study were to investigate the relationship between the levels of histone acetylation (AcH4K5) and mRNA expression level of histone deacetylase 1(HDAC1) during early development of porcine parthenogenetic(PA) embryos in vitro.[Method]Porcine PA embryos (2-cell, 4-cell, morula and blastocyst) were collected, histone acetylation levels of AcH4K5 of four states' embryos were assessed using immunofluorescence and laser confocal microscope by detecting the fluorescence intensity.Real-time PCR method was used for quantitative analysis of HDAC1 mRNA level in PA embryos.[Result]The results showed that AcH4K5 expressed in nucleolus of all embryos from 2-cell to blastocyst, and the levels of AcH4K5 expression increased gradually, reached the highest in blastocyst stage, and the expression at each stage was different significantly(1124.77±127.78, 1305.81±184.23, 1795.19±318.67, and 2149.63±529.47, P＜0.05, respectively).There were dynamic changes of HDAC1 mRNA level among four states'embryos (2-cell, 4-cell, morula and blastocyst of PA embryos), and mRNA expression patterns of HDAC1 genes decreased from 2-cell to blastocyst stage (1.00±0, 0.91±0.009, 0.85±0.008, 0.67±0.006, P＜0.05, respectively).[Conclusion]These results indicated that there is an inverse correlation between the levels of AeH4K5 and mRNA expression of HDAC1 genes from 2-cell to blastocyst.