Department of Cell and Developmental Biology, University of Illinois at Urbana-Champaign , Urbana, IL, USA.
Cell Cycle. 2020 Nov;19(21):2927-2938. doi: 10.1080/15384101.2020.1829823. Epub 2020 Oct 12.
RFWD3 is an E3 ubiquitin ligase that plays important roles in DNA damage response and DNA replication. We have previously demonstrated that the stabilization of RFWD3 by PCNA at the replication fork enables ubiquitination of the single-stranded binding protein, RPA and its subsequent degradation for replication progression. Here, we report that RFWD3 associates with the Origin Recognition Complex (ORC) and ORC-Associated (ORCA/LRWD1), components of the pre-replicative complex required for the initiation of DNA replication. Overexpression of ORC/ORCA leads to the stabilization of RFWD3. Interestingly, RFWD3 seems to stabilize ORC/ORCA in cells expressing wild type p53, as the depletion of RFWD3 reduces the levels of ORC/ORCA. Further, the catalytic activity of RFWD3 is required for the stabilization of ORC. Our results indicate that the RFWD3 promotes the stability of ORC, enabling efficient pre-RC assembly.
RFWD3 是一种 E3 泛素连接酶，在 DNA 损伤反应和 DNA 复制中发挥重要作用。我们之前已经证明，PCNA 在复制叉处稳定 RFWD3，从而使单链结合蛋白 RPA 发生泛素化及其随后的降解，以促进复制进程。在这里，我们报告 RFWD3 与起始复制所需的预复制复合物的组成部分复制原点识别复合物（ORC）和 ORC 相关蛋白（ORCA/LRWD1）相关联。ORC/ORCA 的过表达导致 RFWD3 的稳定。有趣的是，在表达野生型 p53 的细胞中，RFWD3 似乎稳定 ORC/ORCA，因为 RFWD3 的耗竭会降低 ORC/ORCA 的水平。此外，RFWD3 的催化活性对于 ORC 的稳定是必需的。我们的结果表明，RFWD3 促进 ORC 的稳定性，从而实现有效的预 RC 组装。
