Department of Biochemistry and Molecular Biology, Tianjin Medical University, 300070, Tianjin, P. R. China.
Key Laboratory of Immune Microenvironment and Disease (Ministry of Education), Tianjin Medical University, 300070, Tianjin, P. R. China.
Nat Commun. 2021 Jan 12;12(1):321. doi: 10.1038/s41467-020-20538-w.
The yeast protein Rad5 and its orthologs in other eukaryotes promote replication stress tolerance and cell survival using their multiple activities, including ubiquitin ligase, replication fork remodeling and DNA lesion targeting activities. Here, we present the crystal structure of a nearly full-length Rad5 protein. The structure shows three distinct, but well-connected, domains required for Rad5's activities. The spatial arrangement of these domains suggest that different domains can have autonomous activities but also undergo intrinsic coordination. Moreover, our structural, biochemical and cellular studies demonstrate that Rad5's HIRAN domain mediates interactions with the DNA metabolism maestro factor PCNA and contributes to its poly-ubiquitination, binds to DNA and contributes to the Rad5-catalyzed replication fork regression, defining a new type of HIRAN domains with multiple activities. Our work provides a framework to understand how Rad5 integrates its various activities in replication stress tolerance.
酵母蛋白 Rad5 及其在其他真核生物中的同源物通过其多种活性，包括泛素连接酶、复制叉重塑和 DNA 损伤靶向活性，促进复制应激耐受和细胞存活。在这里，我们展示了一个近乎全长 Rad5 蛋白的晶体结构。该结构显示了 Rad5 活性所需的三个截然不同但连接良好的结构域。这些结构域的空间排列表明不同的结构域可以具有自主活性，但也会进行内在协调。此外，我们的结构、生化和细胞研究表明，Rad5 的 HIRAN 结构域介导与 DNA 代谢大师因子 PCNA 的相互作用，并有助于其多泛素化，与 DNA 结合并有助于 Rad5 催化的复制叉回归，定义了具有多种活性的新型 HIRAN 结构域。我们的工作为理解 Rad5 如何整合其在复制应激耐受中的各种活性提供了一个框架。
