Plasmids of incompatibility group P (IncP) show an extraordinary host range among gram-negative bacteria. The genetic and molecular basis for this potential is not known, and we are studying in detail the IncP plasmid RK2 in the belief that it may reveal special genetic functions or unique strategies for gene expression. We have found that RK2 encodes two novel sets of genetic determinants: (1) kil genes (kilA, kilB, kilC) which are lethal to an E, coli host, and (2) kor genes (korB, korC) which control the kil genes. Our studies indicate that these may be encoded by all IncP plasmids, and we have speculated that these may be host range determinants. We have learned that these genes are involved in an array of regulatory interactions, including the control of plasmid regulation. Here we wish to continue our investigations of these genes. Specifically we intend to do the following: (1) to complete the genetic mapping and to determine the nucleotide sequences of each of the kil and kor determinants, (2) to identify and characterize the gene products, (3) to learn how kil functions are lethal to E coli, (4) to determine the molecular mechanisms by which the kor functions prevent cell death by the kil genes, (5) to determine the molecular basis of korA-mediated activation of korB and korC expression, (6) to study the evolutionary relationships of IncP plasmids by hybridization analysis and nucleotide sequence comparisons of their kil and kor determinants, and (7) to test the relationship of host range to kil and kor functions. By employing a variety of both genetic and molecular techniques, we hope to achieve a complete understanding of the kil and kor functions and their importance to the biology of IncP plasmids.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM029085-07
Application #
3276567
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Project Start
1981-04-01
Project End
1989-03-31
Budget Start
1987-04-01
Budget End
1988-03-31
Support Year
7
Fiscal Year
1987
Total Cost
Indirect Cost
Name
Columbia University (N.Y.)
Department
Type
Schools of Medicine
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10027
Siddique, Azeem; Figurski, David H (2012) Different phenotypes of Walker-like A box mutants of ParA homolog IncC of broad-host-range IncP plasmids. Plasmid 68:93-104
Siddique, Azeem; Figurski, David H (2002) The active partition gene incC of IncP plasmids is required for stable maintenance in a broad range of hosts. J Bacteriol 184:1788-93
Kachlany, Scott C; Fine, Daniel H; Figurski, David H (2002) Purification of secreted leukotoxin (LtxA) from Actinobacillus actinomycetemcomitans. Protein Expr Purif 25:465-71
Wilson, James W; Figurski, David H (2002) Host-specific incompatibility by 9-bp direct repeats indicates a role in the maintenance of broad-host-range plasmid RK2. Plasmid 47:216-23
Bhattacharyya, A; Figurski, D H (2001) A small protein-protein interaction domain common to KlcB and global regulators KorA and TrbA of promiscuous IncP plasmids. J Mol Biol 310:51-67
Rosche, T M; Siddique, A; Larsen, M H et al. (2000) Incompatibility protein IncC and global regulator KorB interact in active partition of promiscuous plasmid RK2. J Bacteriol 182:6014-26
Wilson, J W; Sia, E A; Figurski, D H (1997) The kilE locus of promiscuous IncP alpha plasmid RK2 is required for stable maintenance in Pseudomonas aeruginosa. J Bacteriol 179:2339-47
Jovanovic, O S; Figurski, D H (1997) A potential new gene (tccA) on IncP plasmid RK2 and transposon Tn1721: relationship of its product to the TrwC relaxase/helicase of IncW plasmid R388. Plasmid 38:220-3
Sia, E A; Kuehner, D M; Figurski, D H (1996) Mechanism of retrotransfer in conjugation: prior transfer of the conjugative plasmid is required. J Bacteriol 178:1457-64
Sia, E A; Roberts, R C; Easter, C et al. (1995) Different relative importances of the par operons and the effect of conjugal transfer on the maintenance of intact promiscuous plasmid RK2. J Bacteriol 177:2789-97

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