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Interplay between type 1A topoisomerases and gyrase in chromosome segregation in Escherichia coli.

ABSTRACT: Escherichia coli possesses two type 1A topoisomerases, Topo I (topA) and Topo III (topB). Topo I relaxes excess negative supercoiling, and topA mutants can grow only in the presence of compensatory mechanisms, such as gyrase mutations. topB mutants grow as well as wild-type cells. In vitro, Topo III, but not Topo I, can efficiently decatenate DNA during replication. However, in vivo, a chromosome segregation defect is seen only when both type 1A topoisomerases are absent. Here we present experimental evidence for an interplay between gyrase and type 1A topoisomerases in chromosome segregation. We found that both the growth defect and the Par(-) phenotypes of a gyrB(Ts) mutant at nonpermissive temperatures were significantly corrected by deleting topA, but only when topB was present. Overproducing Topo IV, the major cellular decatenase, could not substitute for topB. We also show that overproducing Topo III at a very high level could suppress the Par(-) phenotype. We previously found that the growth and chromosome segregation defects of a triple topA rnhA gyrB(Ts) mutant in which gyrase supercoiling activity was strongly inhibited could be corrected by overproducing Topo III (V. Usongo, F. Nolent, P. Sanscartier, C. Tanguay, S. Broccoli, I. Baaklini, K. Drlica, and M. Drolet, Mol. Microbiol. 69:968-981, 2008). We show here that this overproduction could be bypassed by substituting the gyrB(Ts) allele for a gyrB(+) one or by growing cells in a minimal medium, conditions that reduced both topA- and rnhA-dependent unregulated replication. Altogether, our data point to a role for Topo III in chromosome segregation when gyrase is inefficient and suggest that Topo I plays an indirect role via supercoiling regulation.

SUBMITTER: Usongo V

PROVIDER: S-EPMC3624552 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature

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Interplay between type 1A topoisomerases and gyrase in chromosome segregation in Escherichia coli.

Usongo Valentine V Tanguay Cynthia C Nolent Flora F Bessong Jill Egbe JE Drolet Marc M

Journal of bacteriology 20130208 8

Escherichia coli possesses two type 1A topoisomerases, Topo I (topA) and Topo III (topB). Topo I relaxes excess negative supercoiling, and topA mutants can grow only in the presence of compensatory mechanisms, such as gyrase mutations. topB mutants grow as well as wild-type cells. In vitro, Topo III, but not Topo I, can efficiently decatenate DNA during replication. However, in vivo, a chromosome segregation defect is seen only when both type 1A topoisomerases are absent. Here we present experim ...[more]

PMID: 23396913

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Project description:In eukaryotes, type 1A topoisomerases (topos) act with RecQ-like helicases to maintain the stability of the genome. Despite having been the first type 1A enzymes to be discovered, much less is known about the involvement of the E. coli topo I (topA) and III (topB) enzymes in genome maintenance. These enzymes are thought to have distinct cellular functions: topo I regulates supercoiling and R-loop formation, and topo III is involved in chromosome segregation. To better characterize their roles in genome maintenance, we have used genetic approaches including suppressor screens, combined with microscopy for the examination of cell morphology and nucleoid shape. We show that topA mutants can suffer from growth-inhibitory and supercoiling-dependent chromosome segregation defects. These problems are corrected by deleting recA or recQ but not by deleting recJ or recO, indicating that the RecF pathway is not involved. Rather, our data suggest that RecQ acts with a type 1A topo on RecA-generated recombination intermediates because: 1-topo III overproduction corrects the defects and 2-recQ deletion and topo IIII overproduction are epistatic to recA deletion. The segregation defects are also linked to over-replication, as they are significantly alleviated by an oriC::aph suppressor mutation which is oriC-competent in topA null but not in isogenic topA+ cells. When both topo I and topo III are missing, excess supercoiling triggers growth inhibition that correlates with the formation of extremely long filaments fully packed with unsegregated and diffuse DNA. These phenotypes are likely related to replication from R-loops as they are corrected by overproducing RNase HI or by genetic suppressors of double topA rnhA mutants affecting constitutive stable DNA replication, dnaT::aph and rne::aph, which initiates from R-loops. Thus, bacterial type 1A topos maintain the stability of the genome (i) by preventing over-replication originating from oriC (topo I alone) and R-loops and (ii) by acting with RecQ.

Dataset Information

Interplay between type 1A topoisomerases and gyrase in chromosome segregation in Escherichia coli.

Publications

Interplay between type 1A topoisomerases and gyrase in chromosome segregation in Escherichia coli.

Similar Datasets

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