Moore G D, Ayabe T, Kopf G S, Schultz R M
Department of Obstetrics and Gynecology, University of Pennsylvania, Philadelphia 19104-6018, USA.
Mol Reprod Dev. 1996 Nov;45(3):264-75. doi: 10.1002/(SICI)1098-2795(199611)45:3<264::AID-MRD2>3.0.CO;2-Q.
Cell-cycle progression in somatic cells is regulated by a family of cyclins and cyclindependent kinases (cdks) that form specific complexes as a function of cell-cycle progression. However, the transcript abundance of G1-S cyclins and cdks during the meiotic and mitotic cell cycles of mammalian embryos has not been previously reported. Using a reverse transcription-polymerase chain reaction (PCR) assay that detects changes in either mRNA abundance or polyadenylation state, we examined the relative levels of gene expression for the G1-S cyclins and cdks, as well as for p21, p27, and the retinoblastoma (Rb) gene in mouse oocytes, metaphase II-arrested eggs, and 1-2-cell embryos. The PCR products for cyclins D1, D3, and A, as well as cdk4, p21, and Rb, displayed similar levels in meiotically incompetent and competent oocytes, as well as in metaphase II-arrested eggs. The levels of PCR products for cyclin D2, p27, and two forms of cdk2 were similar in meiotically incompetent and competent oocytes but decreased during oocyte maturation. Finally, the level of PCR products for cyclin E and cdk2 gradually decreased during the progression from meiotically incompetent oocytes to metaphase II-arrested eggs. When the levels of PCR products for the G1-S regulatory genes were evaluated during the first and second mitotic cell cycles, four main patterns were found: 1) steady levels for cyclin A; 2) steady levels followed by a 2-3-fold increase during the G2 phase of the second mitotic cell cycle for cyclins D1, E, cdk2, and p21; 3) a transient increase during the S and/or G2 phases of the first mitotic cell cycle for p27, cyclin D3, and the two forms of cdk2; and 4) higher levels during the first cell cycle and then a decrease with lower levels during the second mitotic cell cycle for cyclin D2 and Rb. cdk4 expression displayed a combination of patterns 2 and 3. The increase in the amount of PCR product for the cdk4 gene during the first mitotic cell cycle was due to polyadenylation, whereas the increase in the amount of PCR product for cdk4, cdk2, and cyclins D1 and E in the second mitotic cell cycle was a product of activation of the embryonic genome.
体细胞中的细胞周期进程受细胞周期蛋白和细胞周期蛋白依赖性激酶（cdks）家族调控，它们会随着细胞周期进程形成特定复合物。然而，此前尚未有关于哺乳动物胚胎减数分裂和有丝分裂细胞周期中G1-S期细胞周期蛋白和cdks转录本丰度的报道。我们使用逆转录-聚合酶链反应（PCR）分析方法来检测mRNA丰度或多聚腺苷酸化状态的变化，研究了小鼠卵母细胞、中期II期阻滞的卵以及1-2细胞胚胎中G1-S期细胞周期蛋白和cdks以及p21、p27和视网膜母细胞瘤（Rb）基因的相对基因表达水平。细胞周期蛋白D1、D3和A以及cdk4、p21和Rb的PCR产物在减数分裂能力不足和有能力的卵母细胞以及中期II期阻滞的卵中显示出相似水平。细胞周期蛋白D2、p27和两种形式的cdk2的PCR产物水平在减数分裂能力不足和有能力的卵母细胞中相似，但在卵母细胞成熟过程中下降。最后，从减数分裂能力不足的卵母细胞到中期II期阻滞的卵的过程中，细胞周期蛋白E和cdk2的PCR产物水平逐渐下降。当在第一次和第二次有丝分裂细胞周期中评估G1-S期调控基因的PCR产物水平时，发现了四种主要模式：1）细胞周期蛋白A水平稳定；2）细胞周期蛋白D1、E、cdk2和p21在第二次有丝分裂细胞周期的G2期水平稳定，随后增加2-3倍；3）p27、细胞周期蛋白D3和两种形式的cdk2在第一次有丝分裂细胞周期的S期和/或G2期短暂增加；4）细胞周期蛋白D2和Rb在第一个细胞周期中水平较高，然后在第二次有丝分裂细胞周期中下降至较低水平。cdk4的表达表现出模式2和模式3的组合。第一次有丝分裂细胞周期中cdk4基因PCR产物量的增加是由于多聚腺苷酸化，而第二次有丝分裂细胞周期中cdk4、cdk2以及细胞周期蛋白D1和E的PCR产物量的增加是胚胎基因组激活的结果。
