Maintenance of Genome Stability and Breast Cancer: Molecular Analysis of DNA Damage-Activated Kinases
Annual summary rept. 15 Feb 2006-17 Feb 2007
VANDERBILT UNIV MEDICAL CENTER NASHVILLE TN
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The ATR ATM and Rad3-Related kinase is essential to maintain genomic integrity. ATR is recruited to DNA lesions in part through its association with ATR-interacting protein ATRIP, which in turn interacts with the single-stranded DNA binding protein RPA Replication Protein A. In this study, a conserved checkpoint protein recruitment domain CRD in ATRIP orthologs has been identified by biochemical mapping of the RPA binding site in combination with NMR, mutagenesis and computational modeling. Mutations in the CRD of the yeast ATRIP ortholog Ddc2 disrupt the Ddc2-RPA interaction, prevent proper localization of Ddc2 to DNA breaks, sensitize yeast to DNA damaging agents, and partially compromise checkpoint signaling. These data demonstrate that the CRD is critical for localization and optimal DNA damage responses. However, the stimulation of ATR kinase activity by binding of TopBP1 to ATRIP-ATR can occur independently of the interaction of ATRIP with RPA. Our results support a multi-step model for ATR activation that requires separable localization and activation functions of ATRIP.
- Genetic Engineering and Molecular Biology
- Medicine and Medical Research