Abstract

The broad objective of this proposal is to understand the biochemical events that comprise the process of genetic recombination. This goal will be achieved by reconstituting various steps of the recombination process in vitro, using purified proteins from Escherichia coli and from Saccharomyces cerevisiae, and also by understanding the mechanism of these molecular events using defined biochemical reactions. Genetic recombination is a fundamental biological process that involves the processing of broken DNA, homologous recognition, exchange of DNA strands, and resolution of the recombination intermediates. It is an important cellular process that is used by all organisms to repair DNA damage, restart DNA replication, and generate genetic diversity. Two specific objectives are planned. The first set of aims is to fully reconstitute in vitro the genetic recombination process of E. coli. The second major aim is to determine the biochemical mechanism of steps that define recombination in the eukaryote, S. cerevisiae. Understanding the mechanism of recombination will provide insight into the manner by which DNA breaks are repaired and, thereby, ensure that genomes are accurately repaired and maintained. Failure to repair DNA properly can ultimately lead to cancer development. The human homologs of some of the proteins that we will investigate include the Breast Cancer Susceptibility Gene 2 (BRCA2) and the Bloom (BLM) helicase. Understanding the mechanism of recombination should also provide insight into possible new experimental approaches for gene replacement therapies.

Public Health Relevance

Recombination is an important cellular process that is used by all organisms to repair DNA damage. Understanding the mechanism of recombination will provide insight into the manner by which DNA breaks are repaired and, thereby, ensure that genomes are accurately repaired and maintained. Failure to repair DNA properly can ultimately lead to cancer development, premature aging, and anemia.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM062653-30
Application #
7889116
Study Section
Molecular Genetics A Study Section (MGA)
Program Officer
Hagan, Ann A
Project Start
1982-04-01
Project End
2014-03-31
Budget Start
2010-04-01
Budget End
2011-03-31
Support Year
30
Fiscal Year
2010
Total Cost
$538,384
Indirect Cost

  Institution

Name
University of California Davis
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
047120084
City
Davis
State
CA
Country
United States
Zip Code
95618

  Publications

Gong, Yi; Handa, Naofumi; Kowalczykowski, Stephen C et al. (2017) PHF11 promotes DSB resection, ATR signaling, and HR. Genes Dev 31:46-58
Martinez, Juan S; von Nicolai, Catharina; Kim, Taeho et al. (2016) BRCA2 regulates DMC1-mediated recombination through the BRC repeats. Proc Natl Acad Sci U S A 113:3515-20
Bell, Jason C; Kowalczykowski, Stephen C (2016) RecA: Regulation and Mechanism of a Molecular Search Engine. Trends Biochem Sci 41:491-507
Pavankumar, T L; Exell, J C; Kowalczykowski, S C (2016) Direct Fluorescent Imaging of Translocation and Unwinding by Individual DNA Helicases. Methods Enzymol 581:1-32
Kowalczykowski, Stephen C (2015) An Overview of the Molecular Mechanisms of Recombinational DNA Repair. Cold Spring Harb Perspect Biol 7:
Fasching, Clare L; Cejka, Petr; Kowalczykowski, Stephen C et al. (2015) Top3-Rmi1 dissolve Rad51-mediated D loops by a topoisomerase-based mechanism. Mol Cell 57:595-606
Chen, Huan; Donnianni, Roberto A; Handa, Naofumi et al. (2015) Sae2 promotes DNA damage resistance by removing the Mre11-Rad50-Xrs2 complex from DNA and attenuating Rad53 signaling. Proc Natl Acad Sci U S A 112:E1880-7
Bell, Jason C; Liu, Bian; Kowalczykowski, Stephen C (2015) Imaging and energetics of single SSB-ssDNA molecules reveal intramolecular condensation and insight into RecOR function. Elife 4:e08646
Wang, Anderson T; Kim, Taeho; Wagner, John E et al. (2015) A Dominant Mutation in Human RAD51 Reveals Its Function in DNA Interstrand Crosslink Repair Independent of Homologous Recombination. Mol Cell 59:478-90
Bocquet, Nicolas; Bizard, Anna H; Abdulrahman, Wassim et al. (2014) Structural and mechanistic insight into Holliday-junction dissolution by topoisomerase III? and RMI1. Nat Struct Mol Biol 21:261-8

Showing the most recent 10 out of 53 publications