Selected amplification of the cell division genes ftsQ-ftsA-ftsZ in Escherichia coli.
Rapidly growing Escherichia coli is unable to divide in the presence of the antibiotic mecillinam, whose direct target is penicillin-binding protein 2 (PBP2), responsible for the elongation of the cylindrical portion of the cell wall. Division can be restored in the absence of PBP2 activity by increasing the concentration of the cell division proteins FtsQ, FtsA, and FtsZ. We tried to identify regulators of the ftsQ-ftsA-ftsZ operon among mecillinam-resistant mutants, which include strains overexpressing these genes. By insertional mutagenesis with mini-Tn10 elements, we selected for insertions that conferred mecillinam resistance. Among 15 such mutants, 7 suppressed the thermosensitivity of the ftsZ84(Ts) mutant, strongly suggesting that they had increased FtsZ activity. In all 7 cases, however, the mutants resulted from a duplication of the ftsQAZ region. These duplications seemed to result from multiple events, suggesting that no simple insertional inactivation can result in a mutant with sufficiently amplified ftsQAZ expression to confer mecillinam resistance. The structure of the duplications suggests a general method for constructing directed duplications of precise sequences.
Vinella
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Institut Jacques Monod (CNRS, Université Paris 7, Université Paris 6), 75251 Paris Cedex 05, France. vinella@ijm.jussieu.fr
Cashel
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M
D'Ari
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R
D000560:Amdinocillin / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics*; D002455:Cell Division / Q000235:genetics; D003598:Cytoskeletal Proteins; D004251:DNA Transposable Elements / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology; D004352:Drug Resistance, Microbial / Q000235:genetics; D004926:Escherichia coli / Q000166:cytology / Q000187:drug effects / Q000235:genetics*; D029968:Escherichia coli Proteins; D005784:Gene Amplification; D015964:Gene Expression Regulation, Bacterial; D006159:Guanosine Tetraphosphate / Q000502:physiology; D006358:Hot Temperature; D008565:Membrane Proteins / Q000235:genetics*; D008957:Models, Genetic; D016254:Mutagenesis, Insertional; D009876:Operon / Q000235:genetics; D015800:Protozoan Proteins; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology; D014161:Transduction, Genetic
D001426:Bacterial Proteins; D003598:Cytoskeletal Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C065103:FtsA protein, Bacteria; C489310:FtsA protein, E coli; C068023:FtsQ protein, E coli; C071525:FtsZ protein, Bacteria; D008565:Membrane Proteins; C400748:PBP-2 protein, Leptomonas seymouri; D015800:Protozoan Proteins; D014157:Transcription Factors; D006159:Guanosine Tetraphosphate; D000560:Amdinocillin