Influence of the Human Cohesion Establishment Factor Ctf4/AND-1 on DNA Replication
Influence of the Human Cohesion Establishment Factor Ctf4/AND-1 on DNA Replication
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DOI：
10.1074/jbc.m109.093609
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发表时间：
2010-03-26
影响因子：
4.8
通讯作者：
Hurwitz, Jerard
中科院分区：
文献类型：
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作者：
Bermudez, Vladimir P.;Farina, Andrea;Hurwitz, Jerard
Ctf 4/AND-1是一个高度保守的基因产物，DNA复制和姐妹染色单体凝聚力的建立都需要它。在这份报告中，我们研究了人Ctf 4（hCtf 4）在体外和体内DNA复制的作用机制。我们的研究结果表明，纯化的hCtf 4作为二聚体存在，并且hCtf 4 SepB结构域可能起着决定二聚体结构的主要作用。hCtf 4优先结合DNA模板-引物结构，直接与复制型DNA聚合酶（α、δ和β）相互作用，并在体外显著刺激DNA聚合酶α和β的聚合酶活性。用小分子干扰RNA（siRNA）去除HeLa细胞中的hCtf 4，可使HeLa细胞发生G（1）/S期阻滞。DNA纤维分析表明，细胞耗尽的hCtf 4表现出的DNA复制速率慢于用对照小干扰RNA处理的细胞。这些发现表明，在人类细胞中，hCtf 4在DNA复制中起着至关重要的作用，其刺激复制性DNA聚合酶的能力可能有助于这种效果。
Ctf4/AND-1 is a highly conserved gene product required for both DNA replication and the establishment of sister chromatid cohesion. In this report, we examined the mechanism of action of human Ctf4 (hCtf4) in DNA replication both in vitro and in vivo. Our findings show that the purified hCtf4 exists as a dimer and that the hCtf4 SepB domain likely plays a primary role determining the dimeric structure. hCtf4 binds preferentially to DNA template-primer structures, interacts directly with the replicative DNA polymerases (alpha, delta, and epsilon), and markedly stimulates the polymerase activities of DNA polymerases alpha and epsilon in vitro. Depletion of hCtf4 in HeLa cells by small interfering RNA resulted in G(1)/S phase arrest. DNA fiber analysis revealed that cells depleted of hCtf4 exhibited a rate of DNA replication slower than cells treated with control small interfering RNA. These findings suggest that in human cells, hCtf4 plays an essential role in DNA replication and its ability to stimulate the replicative DNA polymerases may contribute to this effect.