Overexpression of the Tn5 transposase in Escherichia coli results in filamentation, aberrant nucleoid segregation, and cell death: analysis of E. coli and transposase suppressor mutations.

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RESUMO

Overexpression of the Tn5 transposase (Tnp) was found to be lethal to Escherichia coli. This killing was not caused by transposition or dependent on the transpositional or DNA binding competence of Tnp. Instead, it was strictly correlated with the presence of a wild-type N terminus. Deletions removing just two N-terminal amino acids of Tnp resulted in partial suppression of this effect, and deletions of Tnp removing 3 or 11 N-terminal amino acids abolished the killing effect. This cytotoxic effect of Tnp overexpression is accompanied by extensive filament formation (i.e., a defect in cell division) and aberrant nucleoid segregation. Four E. coli mutants were isolated which allow survival upon Tnp overexpression, and the mutations are located at four discrete loci. These suppressor mutations map near essential genes involved in cell division and DNA segregation. One of these mutations maps to a 4.5-kb HindIII region containing the ftsYEX (cell division) locus at 76 min. A simple proposition which accounts for all of these observations is that Tnp interacts with an essential E. coli factor affecting cell division and/or chromosome segregation and that overexpression of Tnp titrates this factor below a level required for viability of the cell. Furthermore, the N terminus of Tnp is necessary for this interaction. The possible significance of this phenomenon for the transposition process is discussed.

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