Costanzo V, Avvedimento E V, Gottesman M E, Gautier J, Grieco D
Dipartimento di Biologia e Patologia Molecolare e Cellulare "L. Califano", c/o Centro di Endocrinologia ed Oncologia Sperimentale del CNR, II Medical School, University of Naples, Via S. Pansini 5, Naples, 80131, Italy.
Curr Biol. 1999 Aug 26;9(16):903-6. doi: 10.1016/s0960-9822(99)80395-9.
Passage through mitosis resets cells for a new round of chromosomal DNA replication [1]. In late mitosis, the pre-replication complex - which includes the origin recognition complex (ORC), Cdc6 and the minichromosome maintenance (MCM) proteins - binds chromatin as a pre-requisite for DNA replication. S-phase-promoting cyclin-dependent kinases (Cdks) and the kinase Dbf4-Cdc7 then act to initiate replication. Before the onset of replication Cdc6 dissociates from chromatin. S-phase and M-phase Cdks block the formation of a new pre-replication complex, preventing DNA over-replication during the S, G2 and M phases of the cell cycle [1]. The nuclear membrane also contributes to limit genome replication to once per cell cycle [2]. Thus, at the end of M phase, nuclear membrane breakdown and the collapse of Cdk activity reset cells for a new round of chromosomal replication. We showed previously that protein kinase A (PKA) activity oscillates during the cell cycle in Xenopus egg extracts, peaking in late mitosis. The oscillations are induced by the M-phase-promoting Cdk [3] [4]. Here, we found that PKA oscillation was required for the following phase of DNA replication. PKA activity was needed from mitosis exit to the formation of the nuclear envelope. PKA was not required for the assembly of ORC2, Cdc6 and MCM3 onto chromatin. Inhibition of PKA activity, however, blocked the release of Cdc6 from chromatin and subsequent DNA replication. These data suggest that PKA activation in late M phase is required for the following S phase.
有丝分裂过程会使细胞重置，以进行新一轮的染色体DNA复制[1]。在有丝分裂后期，前复制复合体（包括起始识别复合体（ORC）、Cdc6和微小染色体维持（MCM）蛋白）结合染色质，这是DNA复制的前提条件。促进S期的细胞周期蛋白依赖性激酶（Cdks）和激酶Dbf4-Cdc7随后启动复制。在复制开始前，Cdc6从染色质上解离。S期和M期的Cdks会阻止新的前复制复合体形成，从而防止在细胞周期的S期、G2期和M期发生DNA过度复制[1]。核膜也有助于将基因组复制限制在每个细胞周期一次[2]。因此，在M期结束时，核膜破裂和Cdk活性的下降会使细胞重置，以进行新一轮的染色体复制。我们之前发现，在非洲爪蟾卵提取物中，蛋白激酶A（PKA）的活性在细胞周期中振荡，在有丝分裂后期达到峰值。这种振荡是由促进M期的Cdk诱导的[3][4]。在这里，我们发现DNA复制的后续阶段需要PKA振荡。从有丝分裂退出到核膜形成都需要PKA活性。ORC2、Cdc6和MCM3组装到染色质上不需要PKA。然而，抑制PKA活性会阻止Cdc6从染色质上释放以及随后的DNA复制。这些数据表明，M期后期PKA的激活是后续S期所必需的。
