International Centre for Genetic Engineering and Biotechnology, Padriciano 99, 34149 Trieste, Italy.
Mol Cell Biol. 2010 Mar;30(6):1382-96. doi: 10.1128/MCB.01290-09. Epub 2010 Jan 11.
Cellular and biochemical studies support a role for all five human RecQ helicases in DNA replication; however, their specific functions during this process are unclear. Here we investigate the in vivo association of the five human RecQ helicases with three well-characterized human replication origins. We show that only RECQ1 (also called RECQL or RECQL1) and RECQ4 (also called RECQL4) associate with replication origins in a cell cycle-regulated fashion in unperturbed cells. RECQ4 is recruited to origins at late G(1), after ORC and MCM complex assembly, while RECQ1 and additional RECQ4 are loaded at origins at the onset of S phase, when licensed origins begin firing. Both proteins are lost from origins after DNA replication initiation, indicating either disassembly or tracking with the newly formed replisome. Nascent-origin DNA synthesis and the frequency of origin firing are reduced after RECQ1 depletion and, to a greater extent, after RECQ4 depletion. Depletion of RECQ1, though not that of RECQ4, also suppresses replication fork rates in otherwise unperturbed cells. These results indicate that RECQ1 and RECQ4 are integral components of the human replication complex and play distinct roles in DNA replication initiation and replication fork progression in vivo.
细胞和生化研究支持五种人类 RecQ 解旋酶在 DNA 复制中的作用；然而，它们在这个过程中的具体功能尚不清楚。在这里，我们研究了五种人类 RecQ 解旋酶与三个特征明确的人类复制起点之间的体内关联。我们发现，只有 RECQ1（也称为 RECQL 或 RECQL1）和 RECQ4（也称为 RECQL4）以细胞周期调节的方式与未受干扰的细胞中的复制起点相关联。RECQ4 在晚期 G1 时被招募到原点，在 ORC 和 MCM 复合物组装之后，而 RECQ1 和额外的 RECQ4 在 S 期开始时加载到原点，此时许可的原点开始启动。这两种蛋白质在 DNA 复制起始后都从原点丢失，这表明它们要么解体，要么与新形成的复制体一起追踪。RECQ1 耗尽后，起始原点 DNA 合成和原点启动频率降低，而 RECQ4 耗尽后降低得更多。RECQ1 的耗尽，而不是 RECQ4 的耗尽，也会抑制其他未受干扰的细胞中的复制叉速度。这些结果表明，RECQ1 和 RECQ4 是人类复制复合物的组成部分，在体内 DNA 复制起始和复制叉推进中发挥着不同的作用。
