The long-term goal of the proposed research is to understand how cells preserve genome integrity during DNA replication. Specifically, this application focuses on the ATR (ATM and rad3-related) signaling pathway. ATR functions at the apex of a DNA damage and replication stress response pathway that is needed every cell division cycle to promote the complete and accurate replication of the genome. Many cancer cells are highly dependent on ATR function for proliferation and viability because of elevated levels of oncogene-induced replication stress and mutations in other genome maintenance pathways. Thus, ATR may be a useful drug target based on a synthetic lethal approach. In this proposal we test a specific model of how ATR promotes replication fork stabilization and repair, define the consequences of acute ATR inhibition on DNA replication and cell fate outcomes, define and characterize new ATR-regulated proteins that act at damaged replication forks, characterize cancer settings in which ATR inhibition might be useful using synthetic lethality, and test a new model of how ATR is regulated by autophosphorylation. This is a focused proposal aimed at understanding the most important and least understood aspects of ATR function. Specific hypotheses and innovative concepts based on preliminary data are tested using advanced biochemical and genetic approaches. In addition, the aims provide opportunities for unexpected discoveries about the mechanisms that maintain the genome during DNA replication and when ATR pathway inhibitors may be useful in the cancer clinic.

Public Health Relevance

The replication stress response controlled by the ATR kinase is essential to maintain genome integrity and prevent carcinogenesis. ATR pathway inhibitors are currently being developed as anti-cancer agents. This research proposal will define mechanisms by which ATR promotes genome maintenance during DNA replication and identify cellular consequences of acute disruption of this pathway.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
2R01CA102729-11A1
Application #
8628242
Study Section
Cellular Signaling and Regulatory Systems Study Section (CSRS)
Program Officer
Pelroy, Richard
Project Start
2003-07-01
Project End
2019-04-30
Budget Start
2014-05-06
Budget End
2015-04-30
Support Year
11
Fiscal Year
2014
Total Cost
$264,458
Indirect Cost
$95,834
Name
Vanderbilt University Medical Center
Department
Biochemistry
Type
Schools of Medicine
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37212
Frank, Andreas O; Vangamudi, Bhavatarini; Feldkamp, Michael D et al. (2014) Discovery of a potent stapled helix peptide that binds to the 70N domain of replication protein A. J Med Chem 57:2455-61
Mohni, Kareem N; Kavanaugh, Gina M; Cortez, David (2014) ATR pathway inhibition is synthetically lethal in cancer cells with ERCC1 deficiency. Cancer Res 74:2835-45
Sirbu, Bianca M; Cortez, David (2013) DNA damage response: three levels of DNA repair regulation. Cold Spring Harb Perspect Biol 5:a012724
Souza-Fagundes, Elaine M; Frank, Andreas O; Feldkamp, Michael D et al. (2012) A high-throughput fluorescence polarization anisotropy assay for the 70N domain of replication protein A. Anal Biochem 421:742-9
Sirbu, Bianca M; Couch, Frank B; Cortez, David (2012) Monitoring the spatiotemporal dynamics of proteins at replication forks and in assembled chromatin using isolation of proteins on nascent DNA. Nat Protoc 7:594-605
Nam, Edward A; Zhao, Runxiang; Glick, Gloria G et al. (2011) Thr-1989 phosphorylation is a marker of active ataxia telangiectasia-mutated and Rad3-related (ATR) kinase. J Biol Chem 286:28707-14
Nam, Edward A; Zhao, Runxiang; Cortez, David (2011) Analysis of mutations that dissociate G(2) and essential S phase functions of human ataxia telangiectasia-mutated and Rad3-related (ATR) protein kinase. J Biol Chem 286:37320-7
Nam, Edward A; Cortez, David (2011) ATR signalling: more than meeting at the fork. Biochem J 436:527-36
Mohni, Kareem N; Livingston, Christine M; Cortez, David et al. (2010) ATR and ATRIP are recruited to herpes simplex virus type 1 replication compartments even though ATR signaling is disabled. J Virol 84:12152-64
Lovejoy, Courtney A; Xu, Xin; Bansbach, Carol E et al. (2009) Functional genomic screens identify CINP as a genome maintenance protein. Proc Natl Acad Sci U S A 106:19304-9

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