The overall goal of these studies is to extend our knowledge of mutation in stationary-phase E. coli cells. In one system, reversion of an F'- borne lac frameshift mutation occurs in stationary-phase cells by a novel mutational mechanism requiring RecA, RecBCD, and RuvABC, functions involved in recombination and double-strand break-repair. The proposed experiments will determine whether chromosomal mutations occur by this mechanism, and reveal other routes to mutation in stationary-phase cells as well. The experimental design follows: (1) Independent chromosomal insertions of the lac frameshift mutation target into isogenic F- and F strains will be screened for those that revert to Lac+ during stationary-phase. (2) The dependence of the reversion phenotype of such insertions on homologous recombination functions and the SOS response will be determined. Lac+ mutations themselves will be characterized by mapping and sequence analysis. (3) Chromosomal insertions that are permissive for RecBCD-dependent stationary-phase mutation and some that are not will be exactly located on the E. coli chromosome, creating a map of hot and cold sites. (4) Permissive insertions will be used in a genetic screen to identify additional mechanistic and regulatory components of this mutational mechanism. This work will elucidate universal and F'-specific features of stationary-phase mutation and provide new links between recombination, repair, and mutation. Understanding the mechanisms and regulation of recombination-dependent mutation in stationary-phase or stressed cells has important implications for mutations that lead to bacterial resistance to antibiotics and to the origins of cancer tumor progression and resistance to chemotherapeutic drugs.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Postdoctoral Individual National Research Service Award (F32)
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Special Emphasis Panel (ZRG2-SSS-1 (01))
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Tompkins, Laurie
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Baylor College of Medicine
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Gibson, Janet L; Lombardo, Mary-Jane; Aponyi, Ildiko et al. (2015) Atypical Role for PhoU in Mutagenic Break Repair under Stress in Escherichia coli. PLoS One 10:e0123315
Al Mamun, Abu Amar M; Lombardo, Mary-Jane; Shee, Chandan et al. (2012) Identity and function of a large gene network underlying mutagenic repair of DNA breaks. Science 338:1344-8
Gibson, Janet L; Lombardo, Mary-Jane; Thornton, Philip C et al. (2010) The sigma(E) stress response is required for stress-induced mutation and amplification in Escherichia coli. Mol Microbiol 77:415-30
Lombardo, Mary-Jane; Aponyi, Ildiko; Rosenberg, Susan M (2004) General stress response regulator RpoS in adaptive mutation and amplification in Escherichia coli. Genetics 166:669-80
Lombardo, Mary-Jane; Aponyi, Ildiko; Ray, Mellanie P et al. (2003) xni-deficient Escherichia coli are proficient for recombination and multiple pathways of repair. DNA Repair (Amst) 2:1175-83
Bull, H J; Lombardo, M J; Rosenberg, S M (2001) Stationary-phase mutation in the bacterial chromosome: recombination protein and DNA polymerase IV dependence. Proc Natl Acad Sci U S A 98:8334-41
Gumbiner-Russo, L M; Lombardo, M J; Ponder, R G et al. (2001) The TGV transgenic vectors for single-copy gene expression from the Escherichia coli chromosome. Gene 273:97-104
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Lombardo, M J; Rosenberg, S M (2000) radC102 of Escherichia coli is an allele of recG. J Bacteriol 182:6287-91