Billions of base pairs of DNA must be replicated trillions of times during a human lifetime. Adding to the difficulty, replication is challenged by stresses including DNA template lesions, difficult to replicate sequences, and conflicts with transcription. Cells employ multiple repair and signaling responses to replication stress depending on the type, persistence, and location of the problem. We have employed proteomic and genetic approaches to understand how cells overcome replication stress. These analyses identified several new replication stress response proteins including RADX. RADX binds single-stranded DNA and prevents replication fork cleavage by endonucleases. We hypothesize that it regulates the processes of replication fork reversal and fork protection through its single-stranded DNA binding activity. We will test this hypothesis and more generally characterize mechanisms that regulate fork reversal and protection using biochemical and genetic approaches. Since cancer cells have elevated levels of replication stress and many cancer therapeutics work by interfering with DNA replication, these studies will also generate discoveries that may be translated into the cancer clinic.

Public Health Relevance

This application seeks to understand mechanisms of the replication stress response that stabilize forks and maintain genome stability. Failures in these mechanisms cause cancer and also determine the response of cancer cells to cancer therapies. Therefore, completing this project will provide insights into basic genome maintenance mechanisms with significant clinical application.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM116616-06
Application #
10004147
Study Section
Molecular Genetics B Study Section (MGB)
Program Officer
Reddy, Michael K
Project Start
2015-08-15
Project End
2023-07-31
Budget Start
2020-08-01
Budget End
2021-07-31
Support Year
6
Fiscal Year
2020
Total Cost
Indirect Cost
Name
Vanderbilt University Medical Center
Department
Biochemistry
Type
Schools of Medicine
DUNS #
965717143
City
Nashville
State
TN
Country
United States
Zip Code
37203
Mohni, Kareem N; Wessel, Sarah R; Zhao, Runxiang et al. (2018) HMCES Maintains Genome Integrity by Shielding Abasic Sites in Single-Strand DNA. Cell :
Bhat, Kamakoti P; Krishnamoorthy, Archana; Dungrawala, Huzefa et al. (2018) RADX Modulates RAD51 Activity to Control Replication Fork Protection. Cell Rep 24:538-545
Carvajal-Maldonado, Denisse; Byrum, Andrea K; Jackson, Jessica et al. (2018) Perturbing cohesin dynamics drives MRE11 nuclease-dependent replication fork slowing. Nucleic Acids Res :
Bhat, Kamakoti P; Cortez, David (2018) RPA and RAD51: fork reversal, fork protection, and genome stability. Nat Struct Mol Biol 25:446-453
Poole, Lisa A; Cortez, David (2017) Functions of SMARCAL1, ZRANB3, and HLTF in maintaining genome stability. Crit Rev Biochem Mol Biol 52:696-714
Dungrawala, Huzefa; Bhat, Kamakoti P; Le Meur, Rémy et al. (2017) RADX Promotes Genome Stability and Modulates Chemosensitivity by Regulating RAD51 at Replication Forks. Mol Cell 67:374-386.e5
Vujanovic, Marko; Krietsch, Jana; Raso, Maria Chiara et al. (2017) Replication Fork Slowing and Reversal upon DNA Damage Require PCNA Polyubiquitination and ZRANB3 DNA Translocase Activity. Mol Cell 67:882-890.e5
Reynolds, John J; Bicknell, Louise S; Carroll, Paula et al. (2017) Mutations in DONSON disrupt replication fork stability and cause microcephalic dwarfism. Nat Genet 49:537-549
Cortez, David (2017) Proteomic Analyses of the Eukaryotic Replication Machinery. Methods Enzymol 591:33-53
Poole, Lisa A; Cortez, David (2016) SMARCAL1 and telomeres: Replicating the troublesome ends. Nucleus 7:270-4

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