Dual genome-wide CRISPR knockout and CRISPR activation screens identify  mechanisms that regulate the resistance to multiple ATR inhibitors

by Emily M. Schleicher, Ashna Dhoonmoon, Lindsey M. Jackson, Kristen E. Clements, Coryn L. Stump, Claudia M. Nicolae, George-Lucian Moldovan The ataxia telangiectasia and Rad3-related (ATR) protein kinase is a key regulator of the cellular response to DNA damage. Due to increased amount of replication stress, cancer cells heavily rely on ATR to complete DNA replication and cell cycle progression. Thus, ATR inhibition is an emerging tar get in cancer therapy, with multiple ATR inhibitors currently undergoing clinical trials. Here, we describe dual genome-wide CRISPR knockout and CRISPR activation screens employed to comprehensively identify genes that regulate the cellular resistance to ATR inhibitors. Specifically, we investigated two different ATR inhibitors, namely VE822 and AZD6738, in both HeLa and MCF10A cells. We identified and validated multiple genes that alter the resistance to ATR inhibitors. Importantly, we show that the mechanisms of resistance employed by these genes are varied, and include restoring DNA replica tion fork progression, and prevention of ATR inhibitor-induced apoptosis. In particular, we describe a role for MED12-mediated inhibition of the TGFβ signaling pathway in regulating replication fork stability and cellular survival upon ATR inhibition. Our dual genome-wide screen findings pave the w ay for personalized medicine by identifying potential biomarkers for ATR inhibitor resistance.
Source: PLoS Genetics - Category: Genetics & Stem Cells Authors: Source Type: research