Changes in histone H3 lysine 36 methylation in porcine oocytes and preimplantation embryos.
Changes in histone H3 lysine 36 methylation in porcine oocytes and preimplantation embryos.
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DOI：
10.1371/journal.pone.0100205
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Jin DI
中科院分区：
文献类型：
--
作者：
Diao YF;Oqani RK;Li XX;Lin T;Kang JW;Jin DI
已知组蛋白H3赖氨酸36（H3 K36）甲基化与转录活性基因相关，并且被认为是活性基因座的基因组标记。为了研究H3 K36甲基化在猪中的变化，我们使用特异性抗体和共聚焦显微镜通过免疫细胞化学方法测定了猪胎儿成纤维细胞、卵母细胞和植入前胚胎中H3 K36的单甲基化、双甲基化和三甲基化（分别为H3 K36 me 1、H3 K36 me 2和H3 K36 me 3）。这些分析表明，只有H3 K36 me 3在猪胎儿成纤维细胞一致共定位的转录位点鉴定为积极合成RNA的基础上氟尿苷（FU）的掺入。用阻断转录延伸的flavopiridol处理细胞，完全废除了H3 K36 me 3信号和RNA合成。所有三种类型的H3 K36甲基化都存在，并且在卵母细胞成熟过程中没有显著差异。在孤雌胚胎中，H3 K36 me 1和-me 2在1-细胞至囊胚期胚胎中均有表达。相反，H3 K36 me 3在大多数1-细胞期胚胎中未检测到。H3 K36 me 3信号在2-细胞期胚胎中变得可检测，在4-细胞期达到峰值，在8-细胞期降低，然后在孤雌生殖和体外受精（IVF）胚胎的囊胚期变得不可检测。与体外受精胚胎的情况不同，H3 K36 me 3在体细胞核移植（SCNT）胚胎的1-细胞期不能完全去甲基化。这些结果共同表明，H3 K36 me 3，而不是H3 K36 me 1或-me 2，与猪胎儿成纤维细胞中的转录延伸。H3 K36 me 3在猪胚胎发育过程中受到调控，可能是猪胚胎基因激活的一个组蛋白标记。在SCNT胚胎的核重编程过程中发生了异常的H3 K36三甲基化。
Histone H3 lysine 36 (H3K36) methylation is known to be associated with transcriptionally active genes, and is considered a genomic marker of active loci. To investigate the changes in H3K36 methylation in pig, we determined the mono-, di-, and tri-methylations of H3K36 (H3K36me1, H3K36me2 and H3K36me3, respectively) in porcine fetal fibroblasts, oocytes and preimplantation embryos by immunocytochemistry using specific antibodies and confocal microscopy. These analyses revealed that only H3K36me3 in porcine fetal fibroblasts consistently colocalized with transcription sites identified as actively synthesizing RNA based on fluorouridine (FU) incorporation. Treatment of cells with flavopiridol, which blocks transcription elongation, completely abrogated both H3K36me3 signals and RNA synthesis. All three types of H3K36 methylation were present and did not significantly differ during oocyte maturation. In parthenogenetic embryos, H3K36me1 and -me2 were detected in 1-cell through blastocyst-stage embryos. In contrast, H3K36me3 was not detected in most 1-cell stage embryos. H3K36me3 signals became detectable in 2-cell stage embryos, peaked at the 4-cell stage, decreased at the 8-cell stage, and then became undetectable at blastocyst stages in both parthenogenetic and in vitro-fertilized (IVF) embryos. Unlike the case in IVF embryos, H3K36me3 could not be demethylated completely during the 1-cell stage in somatic cell nuclear transfer (SCNT) embryos. These results collectively indicate that H3K36me3, but not H3K36me1 or -me2, is associated with transcription elongation in porcine fetal fibroblasts. H3K36me3 is developmentally regulated and may be a histone mark of embryonic gene activation in pig. Aberrant H3K36 tri-methylation occurred during the nuclear reprogramming of SCNT embryos.
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