Recombination Repair Complex in Human Cells Widespread genomic instability is a hallmark of human tumors. The most frequent manifestations of genomic instability are large-scale chromosomal rearrangements. Such chromosomal rearrangements can lead to Loss-of-Heterozygosity (LOH) or novel gene fusions encompassing chromosome regions known to contain oncogenes and/or tumor suppressor genes. Recombination repair between partially homologous sequences (homeologous recombination or single-stranded annealing) is a common DMA signature of many of these genomic rearrangement events and is normally suppressed by the mismatch repair machinery. The long-term goal of this research is to understand the biophysical and molecular processes that lead to chromosomal aberrations during human tumorigenesis and their contribution to therapeutic drug resistance. In the last granting period we detailed unique biochemical functions of the fundamental recombination proteins hRAD51, hXRCC2 and hRAD51D as well as both the limitations and surprises to the biochemical role(s) of hMSH2-hMSH6 and hMSH4-hMSH5 in recombination In addition we detailed the individual physical interaction of these DMA repair proteins with numerous other DMA repair components that make up a very large recombination repair complex (Proteome). While the significance of our biophysical studies has been readily apparent, the physiological relevance of complex protein interaction has remained enigmatic. Our biophysical studies have engendered the concept of an ATP-binding protein (A-protein) molecular switch that controls protein interaction(s) and/or function(s). A number of important questions have arisen regarding molecular switch function(s) of the human RecA homplogs (hRAD51, hXRCC2, hRAD51D) when compared with the well-studied prototypical bacterial RecA protein. In this renewal grant application we propose to enhance our understanding of recombination repair in human cells via: I.) Biophysical characterization of the RAD51-paralog molecular switch, II.) Biophysical analysis of hRAD51-paralog activities with histone-DNA substrates, III.) Establish the MMR mechanism(s) that deters homeologous (HOER) and/or single-stranded annealing (SSA) recombination events initiated by RR, and IV.) Examination of the human RR Proteome in vivo and in vitro. It is likely that these studies will significantly contribute to understanding the processes involved in maintaining human chromosome stability and should provide a framework for the design of more efficacious therapeutic methodologies as well as preventive therapeutic modalities designed to control the chromosome instability found in human tumors ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM080176-15
Application #
7254947
Study Section
Radiation Therapeutics and Biology Study Section (RTB)
Program Officer
Portnoy, Matthew
Project Start
1993-03-15
Project End
2010-06-30
Budget Start
2007-07-01
Budget End
2008-06-30
Support Year
15
Fiscal Year
2007
Total Cost
$383,244
Indirect Cost
Name
Ohio State University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
832127323
City
Columbus
State
OH
Country
United States
Zip Code
43210
Senavirathne, Gayan; Lopez Jr, Miguel A; Messer, Ryan et al. (2018) Expression and purification of nuclease-free protocatechuate 3,4-dioxygenase for prolonged single-molecule fluorescence imaging. Anal Biochem 556:78-84
Kar, Anirban; Jones, Nathan; Arat, N Özlem et al. (2018) Long repeating (TTAGGG) n single-stranded DNA self-condenses into compact beaded filaments stabilized by G-quadruplex formation. J Biol Chem 293:9473-9485
Senavirathne, Gayan; Mahto, Santosh K; Hanne, Jeungphill et al. (2017) Dynamic unwrapping of nucleosomes by HsRAD51 that includes sliding and rotational motion of histone octamers. Nucleic Acids Res 45:685-698
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Senavirathne, Gayan; Liu, Jiaquan; Lopez Jr, Miguel A et al. (2015) Widespread nuclease contamination in commonly used oxygen-scavenging systems. Nat Methods 12:901-2
North, Justin A; Amunugama, Ravindra; Klajner, Marcelina et al. (2013) ATP-dependent nucleosome unwrapping catalyzed by human RAD51. Nucleic Acids Res 41:7302-12
Amunugama, Ravindra; Groden, Joanna; Fishel, Richard (2013) The HsRAD51B-HsRAD51C stabilizes the HsRAD51 nucleoprotein filament. DNA Repair (Amst) 12:723-32

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