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bioRxiv · 10.1101/2022.02.04.479194

A novel 5' DNA binding site in RFC facilitates PCNA loading for gap DNA repair

Zhang, F.·Georgescue, R.·Yao, N. Y.·Li, H.·O'Donnell, M. E.
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Abstract

RFC uses ATP to assemble PCNA onto primed sites for replicative DNA polymerases {delta} and {varepsilon}. The RFC pentamer forms a central chamber that binds 3 ss/ds DNA junctions to load PCNA onto DNA during replication. We show here five structures that identify a 2nd DNA binding site in RFC that binds a 5 duplex. This 5 DNA site is located between the N-terminal BRCT domain and AAA+ module of the large Rfc1 subunit. Our structures reveal ideal binding to a 7-nt gap, which includes 2 bp unwound by the clamp loader. Biochemical studies show enhanced binding to 5 and 10 nt gaps, consistent with the structural results. Because both 3 and 5 ends are present at a ssDNA gap, we propose that the 5 site facilitates RFCs PCNA loading activity at a DNA damage-induced gap to recruit gap-filling polymerases. These findings are consistent with genetic studies showing that base excision repair of gaps greater than 1 base requires PCNA and involves the 5 DNA binding domain of Rfc1. We further observe that a 5 end facilitates PCNA loading at an RPA coated 30-nt gap, suggesting a potential role of the RFC 5-DNA site in lagging strand DNA synthesis

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Zhang, F., Georgescue, R., Yao, N. Y., Li, H., O'Donnell, M. E.. 2022-02-05. A novel 5' DNA binding site in RFC facilitates PCNA loading for gap DNA repair. https://doi.org/10.1101/2022.02.04.479194

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