The long term goal of this research is to understand the role of the umuDC gene products of Escherichia coli in the molecular mechanism of induced mutagenesis and in the control of the cell cycle in response to DNA damage. UmuD and UmuC have been shown to permit DNA polymerase III to carry out translesion synthesis which results in mutations (44). Recent results from the Walker laboratory have shown that intact UmuD (the form inactive in translesion synthesis) together with UmuC participate in a cell cycle checkpoint mechanism in response to DNA damage (Opperman, T., S. Murli, and G. C. Walker, manuscript in preparation). Preliminary results from the Walker laboratory suggest that UmuD and UmuD interact physically with the epsilon and beta subunits of DNA polymerase III. Experiments outlined in this proposal are designed to provide insights into how the interaction of UmuD, UmuD and UmuC with these two subunits of DNA polymerase III, and with other cellular proteins is related to their roles in SOS mutagenesis and cell cycle checkpoint control. I will biochemically characterize protein- protein interactions using a variety of techniques including gel filtration and affinity chromatography, a co-immunoprecipitation, chemical cross-linking, and surface plasmon resonance (BIAcore). Subsequent experiments will concentrate on obtaining biochemical evidence of the UmuD and UmuC checkpoint control in vitro.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32CA079161-03
Application #
6174345
Study Section
Special Emphasis Panel (ZRG2-SSS-1 (01))
Program Officer
Lohrey, Nancy
Project Start
2000-08-01
Project End
Budget Start
2000-08-01
Budget End
2001-07-31
Support Year
3
Fiscal Year
2000
Total Cost
$40,936
Indirect Cost
Name
Massachusetts Institute of Technology
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
City
Cambridge
State
MA
Country
United States
Zip Code
02139
Sutton, Mark D; Guzzo, Angelina; Narumi, Issay et al. (2002) A model for the structure of the Escherichia coli SOS-regulated UmuD2 protein. DNA Repair (Amst) 1:77-93
Sutton, Mark D; Narumi, Issay; Walker, Graham C (2002) Posttranslational modification of the umuD-encoded subunit of Escherichia coli DNA polymerase V regulates its interactions with the beta processivity clamp. Proc Natl Acad Sci U S A 99:5307-12
Sutton, M D; Walker, G C (2001) umuDC-mediated cold sensitivity is a manifestation of functions of the UmuD(2)C complex involved in a DNA damage checkpoint control. J Bacteriol 183:1215-24
Sutton, M D; Farrow, M F; Burton, B M et al. (2001) Genetic interactions between the Escherichia coli umuDC gene products and the beta processivity clamp of the replicative DNA polymerase. J Bacteriol 183:2897-909
Sutton, M D; Kim, M; Walker, G C (2001) Genetic and biochemical characterization of a novel umuD mutation: insights into a mechanism for UmuD self-cleavage. J Bacteriol 183:347-57
Sutton, M D; Murli, S; Opperman, T et al. (2001) umuDC-dnaQ Interaction and its implications for cell cycle regulation and SOS mutagenesis in Escherichia coli. J Bacteriol 183:1085-9
Sutton, M D; Walker, G C (2001) Managing DNA polymerases: coordinating DNA replication, DNA repair, and DNA recombination. Proc Natl Acad Sci U S A 98:8342-9
Sutton, M D; Opperman, T; Walker, G C (1999) The Escherichia coli SOS mutagenesis proteins UmuD and UmuD' interact physically with the replicative DNA polymerase. Proc Natl Acad Sci U S A 96:12373-8