Borsuk Ewa, Czolowska Renata
Department of Embryology, Institute of Zoology, Faculty of Biology, University of Warsaw, Poland.
Int J Dev Biol. 2010;54(1):21-31. doi: 10.1387/ijdb.092873eb.
Mammalian primary oocytes are arrested in the post-replicative G2 phase of the cell cycle. In contrast to other G2 nuclei, the nucleus of the growing mouse oocyte can reinitiate DNA synthesis after transfer by cell fusion under favorable cytoplasmic conditions, created by the parthenogenetic one-cell embryo. In the present study, we used the cell hybrid system to analyze the distribution of proteins involved in DNA re-replication in the oocyte nucleus. We show that this process is preceded by an extensive rearrangement of the insoluble fractions of minichromosome maintenance (MCM) proteins (Mcm2, -6 and 7). We also demonstrate that Cdc6 protein is present in primary growing mouse oocytes freshly isolated from the ovary, in a soluble and insoluble form. In contrast to MCM proteins, the insoluble fraction of Cdc6 was not rearranged in oocyte nuclei reinitiating DNA replication in hybrid cells. The rearrangement of MCM proteins and reinitiation of DNA synthesis occurred in the nuclei, in which the nuclear envelope remained intact. Reinitiation of DNA replication in the oocyte nucleus was sensitive to the inhibition of both CDK activity and polyadenylation of maternal mRNAs, indicating a role of proteins synthesized de novo by the embryo. These results allow us to understand better the mechanisms involved in the reinitiation of DNA replication in growing oocytes.
哺乳动物的初级卵母细胞停滞于细胞周期的复制后G2期。与其他G2期细胞核不同，在孤雌单细胞胚胎创造的有利细胞质条件下，通过细胞融合转移后，生长中的小鼠卵母细胞核能够重新启动DNA合成。在本研究中，我们利用细胞杂交系统分析了参与卵母细胞核中DNA重新复制的蛋白质分布。我们发现，这一过程之前伴随着微小染色体维持（MCM）蛋白（Mcm2、-6和7）不溶性部分的广泛重排。我们还证明，Cdc6蛋白以可溶和不溶形式存在于从卵巢新鲜分离的初级生长小鼠卵母细胞中。与MCM蛋白不同，在杂交细胞中重新启动DNA复制的卵母细胞核中，Cdc6的不溶性部分没有重排。MCM蛋白的重排和DNA合成的重新启动发生在核膜保持完整的细胞核中。卵母细胞核中DNA复制的重新启动对CDK活性抑制和母体mRNA的多聚腺苷酸化均敏感，这表明胚胎新合成的蛋白质发挥了作用。这些结果使我们能够更好地理解生长中的卵母细胞中DNA复制重新启动所涉及的机制。
