The Atad5 RFC-like complex is the major unloader of proliferating cell nuclear antigen in Xenopus egg extracts

Kawasoe, Yoshitaka, Shimokawa, Sakiko, Gillespie, Peter J., Blow, J. Julian, Tsurimoto, Toshiki and Takahashi, Tatsuro S. (2024) The Atad5 RFC-like complex is the major unloader of proliferating cell nuclear antigen in Xenopus egg extracts. Journal of Biological Chemistry, 300 (1). ISSN 0021-9258

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Proliferating cell nuclear antigen (PCNA) is a homo-trimeric clamp complex that serves as the molecular hub for various DNA transactions, including DNA synthesis and post-replicative mismatch repair. Its timely loading and unloading are critical for genome stability. PCNA loading is catalyzed by Replication factor C (RFC) and the Ctf18 RFC-like complex (Ctf18-RLC), and its unloading is catalyzed by Atad5/Elg1-RLC. However, RFC, Ctf18-RLC, and even some subcomplexes of their shared subunits are capable of unloading PCNA in vitro, leaving an ambiguity in the division of labor in eukaryotic clamp dynamics. By using a system that specifically detects PCNA unloading, we show here that Atad5-RLC, which accounts for only approximately 3% of RFC/RLCs, nevertheless provides the major PCNA unloading activity in Xenopus egg extracts. RFC and Ctf18-RLC each account for approximately 40% of RFC/RLCs, while immunodepletion of neither Rfc1 nor Ctf18 detectably affects the rate of PCNA unloading in our system. PCNA unloading is dependent on the ATP-binding motif of Atad5, independent of nicks on DNA and chromatin assembly, and inhibited effectively by PCNA-interacting peptides. These results support a model in which Atad5-RLC preferentially unloads DNA-bound PCNA molecules that are free from their interactors.

Item Type: Article
Additional Information: Funding information: This work was supported by JSPS KAKENHI grants [17H06935, 19K16042 to Y. K., and 22H04697, 20H05392, 20H03186 to T. S. T.] and Wellcome Trust Senior Investigator Award WT096598MA to J. J. B.
Uncontrolled Keywords: dna replication,xenopus egg extract,clamp loader,mismatch repair,proliferating cell nuclear antigen,replication clamp
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Depositing User: LivePure Connector
Date Deposited: 10 Jun 2024 11:30
Last Modified: 10 Jun 2024 14:30
DOI: 10.1016/j.jbc.2023.105588


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