Bui Hong-Thuy, Van Thuan Nguyen, Wakayama Teruhiko, Miyano Takashi
Department of Life Science, Graduate School of Science and Technology, Kobe University, Nada-ku, Japan.
Reproduction. 2006 Jun;131(6):1037-49. doi: 10.1530/rep.1.00897.
We examined the involvement of histone H3 modifications in the chromosome condensation and decondensation of somatic cell nuclei injected into mature pig oocytes. Nuclei of pig granulosa cells were transferred into in vitro matured intact pig oocytes, and histone H3 phosphorylation, acetylation, and methylation were examined by immunostaining with specific antibodies in relation to changes in chromosome morphology. In the condensed chromosomes of pig oocytes at metaphase II, histone H3 was phosphorylated at serine 10 (H3-S10) and serine 28 (H3-S28), and methylated at lysine 9 (H3-K9), but was not acetylated at lysine 9, 14 and 18 (H3-K9, H3-K14 and H3-K18). During the first 2 h after nuclear transfer, a series of events were observed in the somatic nuclei: nuclear membrane disassembly; chromosome condensation to form a metaphase-like configuration; an increase in histone H3 phosphorylation levels (H3-S10 and H3-S28). Next, pig oocytes injected with nuclei of somatic cells were electroactivated and the chromosome morphology of oocytes and somatic cells was examined along with histone modifications. Generally, chromosomes of the somatic cells showed a similar progression of cell cycle stage to that of oocytes, through anaphase II- and telophase II-like stages then formed pronucleus-like structures, although the morphology of the spindles differed from that of oocyte spindles. The chromosomes of somatic cells also showed changes in histone H3 dephosphorylation and reacetylation, similar to oocytes. In contrast, histone H3 methylation (H3-K9) of somatic cell nuclei did not show any significant change after injection and electroactivation of the oocytes. These results suggest that nuclear remodeling including histone H3 phosphorylation and acetylation of injected somatic nuclei took place in the oocytes under regulation by the oocyte cytoplasm.
我们研究了组蛋白H3修饰在注入成熟猪卵母细胞的体细胞细胞核染色体凝聚和解凝聚过程中的作用。将猪颗粒细胞的细胞核转移到体外成熟的完整猪卵母细胞中，通过用特异性抗体进行免疫染色，检测组蛋白H3的磷酸化、乙酰化和甲基化与染色体形态变化的关系。在处于减数第二次分裂中期的猪卵母细胞的凝聚染色体中，组蛋白H3在丝氨酸10（H3-S10）和丝氨酸28（H3-S28）位点发生磷酸化，在赖氨酸9（H3-K9）位点发生甲基化，但在赖氨酸9、14和18（H3-K9、H3-K14和H3-K18）位点未发生乙酰化。在核移植后的最初2小时内，在体细胞核中观察到一系列事件：核膜解体；染色体凝聚形成类似中期的构型；组蛋白H3磷酸化水平（H3-S10和H3-S28）增加。接下来，对注射了体细胞细胞核的猪卵母细胞进行电激活，并检查卵母细胞和体细胞的染色体形态以及组蛋白修饰。一般来说，体细胞的染色体显示出与卵母细胞相似的细胞周期阶段进展，经过类似减数第二次分裂后期和末期的阶段，然后形成类原核结构，尽管纺锤体的形态与卵母细胞纺锤体不同。体细胞的染色体也显示出与卵母细胞类似的组蛋白H3去磷酸化和重新乙酰化变化。相比之下，卵母细胞注射和电激活后，体细胞核的组蛋白H3甲基化（H3-K9）没有显示出任何显著变化。这些结果表明，在卵母细胞细胞质的调控下，注入的体细胞核发生了包括组蛋白H3磷酸化和乙酰化在内的核重塑。
