The effect of replication protein A inhibition and post-translational modification on ATR kinase signaling.

Autor: Jordan MR; Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, 64202, USA., Oakley GG; Department of Oral Biology, University of Nebraska Medical Center, Lincoln, NE, 68583, USA., Mayo LD; Department of Pediatrics, Indiana University School of Medicine, Indianapolis, IN, 46202, USA., Balakrishnan L; Department of Biology, School of Science, Indiana University Indianapolis, Indianapolis, IN, 46202, USA., Turchi JJ; Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, 64202, USA. jturchi@iu.edu.; NERx Biosciences Inc., Indianapolis, IN, 46202, USA. jturchi@iu.edu.
Jazyk: angličtina
Zdroj: Scientific reports [Sci Rep] 2024 Aug 26; Vol. 14 (1), pp. 19791. Date of Electronic Publication: 2024 Aug 26.
DOI: 10.1038/s41598-024-70589-y
Abstrakt: The ATR kinase responds to elevated levels of single-stranded DNA (ssDNA) to activate the G2/M checkpoint, regulate origin utilization, preserve fork stability, and allow DNA repair to ensure genome integrity. The intrinsic replication stress in cancer cells makes this pathway an attractive therapeutic target. The ssDNA that drives ATR signaling is sensed by the ssDNA-binding protein replication protein A (RPA), which acts as a platform for ATRIP recruitment and subsequent ATR activation by TopBP1. We have developed chemical RPA inhibitors (RPAi) that block RPA-ssDNA interactions (RPA-DBi) and RPA protein-protein interactions (RPA-PPIi); both activities are required for ATR activation. Here, we biochemically reconstitute the ATR kinase signaling pathway and demonstrate that RPA-DBi and RPA-PPIi abrogate ATR-dependent phosphorylation of target proteins with selectivity advantages over active site ATR inhibitors. We demonstrate that RPA post-translational modifications (PTMs) impact ATR kinase activation but do not alter sensitivity to RPAi. Specifically, phosphorylation of RPA32 and TopBP1 stimulate, while RPA70 acetylation does not affect ATR phosphorylation of target proteins. Collectively, this work reveals the RPAi mechanism of action to inhibit ATR signaling that can be regulated by RPA PTMs and offers insight into the anti-cancer activity of ATR pathway-targeted cancer therapeutics.
(© 2024. The Author(s).)
Databáze: MEDLINE
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