Distinct populations of human PCNA are required for initiation of chromosomal DNA replication and concurrent DNA repair
Distinct populations of human PCNA are required for initiation of chromosomal DNA replication and concurrent DNA repair
复制标题
DOI：
10.1016/j.yexcr.2005.09.009
复制
发表时间：
2005-12-10
影响因子：
3.7
通讯作者：
Krude, T
中科院分区：
文献类型：
--
作者：
Szüts, D;Christov, C;Krude, T
在真核细胞的细胞分裂周期中，基因组DNA的完整性通过调节染色体DNA复制和修复受损DNA来维持。我们已经使用分级分离和重建实验，以纯化人细胞中的G1期晚期模板核中启动人染色体DNA复制的必需因子。在这里，我们报告的识别可溶性PCNA作为一个重要的启动因子在这个系统中。重组组氨酸标记的人PCNA可以替代纯化的内源性人PCNA启动人染色体DNA复制。它特异性地募集到体外起始期间形成的离散DNA复制灶。模板细胞核也含有DNA断裂作为同步程序的结果。一个单独的群体染色质结合的PCNA已经存在于这些模板核中的离散的DNA损伤灶，共定位与γ-H2 AX，RPA和Rad 51。这种DNA损伤相关的PCNA群体以单泛素化为标志，表明它参与DNA修复。重要的是，损伤灶相关的PCNA群体既不参与也不需要在相同的细胞核中启动染色体DNA复制。(c)2005年爱思唯尔公司All rights reserved.
The integrity of genomic DNA during the cell division cycle in eukaryotic cells is maintained by regulated chromosomal DNA replication and repair of damaged DNA. We have used fractionation and reconstitution experiments to purify essential factors for the initiation of human chromosomal DNA replication in late G1 phase template nuclei from human cells. Here, we report the identification of soluble PCNA as an essential initiation factor in this system. Recombinant histidine-tagged human PCNA can substitute for purified endogenous human PCNA to initiate human chromosomal DNA replication. It is recruited specifically to discrete DNA replication foci formed during initiation in vitro. The template nuclei also contain DNA breaks as result of the synchronisation procedure. A separate population of chromatin-bound PCNA is already present in these template nuclei at discrete DNA damage foci, co-localising with gamma-H2AX, RPA and Rad51. This DNA damage-associated PCNA population is marked by mono-ubiquitination, suggesting that it is involved in DNA repair. Importantly, the population of damage focus-associated PCNA is neither involved in, nor required for, the initiation of chromosomal DNA replication in the same nuclei. (c) 2005 Elsevier Inc. All rights reserved.