Ross Pablo J, Ragina Neli P, Rodriguez Ramon M, Iager Amy E, Siripattarapravat Kannika, Lopez-Corrales Nestor, Cibelli Jose B
Departments of, Animal Science Physiology, Michigan State University, East Lansing, Michigan 48824, USA.
Reproduction. 2008 Dec;136(6):777-85. doi: 10.1530/REP-08-0045. Epub 2008 Sep 10.
Trimethylation of histone H3 at lysine 27 (H3K27me3) is established by polycomb group genes and is associated with stable and heritable gene silencing. The aim of this study was to characterize the expression of polycomb genes and the dynamics of H3K27me3 during bovine oocyte maturation and preimplantation development. Oocytes and in vitro-produced embryos were collected at different stages of development. Polycomb gene expression was analyzed by real-time quantitative RT-PCR and immunofluorescence. Global H3K27me3 levels were determined by semiquantitative immunofluorescence. Transcripts for EZH2, EED, and SUZ12 were detected at all stages analyzed, with EZH2 levels being the highest of the three at early stages of development. By the time the embryo reached the blastocyst stage, the level of PcG gene mRNA levels significantly increased. Immunofluorescence staining indicated nuclear expression of EZH2 at all stages while nuclear localized EED and SUZ12 were only evident at the morula and blastocyst stages. Semiquantitative analysis of H3K27me3 levels showed that nuclear fluorescence intensity was the highest in immature oocytes, which steadily decreased after fertilization to reach a nadir at the eight-cell stage, and then increased at the blastocyst stage. These results suggest that the absence of polycomb repressive complex 2 proteins localized to the nucleus of early embryos could be responsible for the gradual decrease in H3K27me3 during early preimplantation development.
组蛋白H3赖氨酸27位点的三甲基化（H3K27me3）由多梳蛋白家族基因建立，并与稳定且可遗传的基因沉默相关。本研究的目的是表征多梳蛋白基因在牛卵母细胞成熟和植入前发育过程中的表达以及H3K27me3的动态变化。在不同发育阶段收集卵母细胞和体外生产的胚胎。通过实时定量RT-PCR和免疫荧光分析多梳蛋白基因的表达。通过半定量免疫荧光测定整体H3K27me3水平。在所有分析阶段均检测到EZH2、EED和SUZ12的转录本，其中EZH2在发育早期的水平是三者中最高的。当胚胎发育到囊胚阶段时，多梳蛋白基因（PcG）的mRNA水平显著增加。免疫荧光染色表明EZH2在所有阶段均有核表达，而核定位的EED和SUZ12仅在桑椹胚和囊胚阶段明显。H3K27me3水平的半定量分析表明，未成熟卵母细胞的核荧光强度最高，受精后其稳步下降，在八细胞阶段达到最低点，然后在囊胚阶段增加。这些结果表明，早期胚胎细胞核中缺乏定位的多梳抑制复合物2蛋白可能是植入前早期发育过程中H3K27me3逐渐减少的原因。
