IS1 transposition is enhanced by translation errors and by bacterial growth at extreme glucose levels.

Transposition of insertion sequences (IS) is an enzyme-mediated process that only occurs in a minority of cells within a bacterial culture. Transposition is thus a rare event, but transposition frequency may vary depending on experimental conditions. For instance in a rich broth, IS elements are known to transpose during stationary phase but not during exponential growth. Using a reporter system which involves the activation of the cryptic bgl operon in Escherichia coli, we show that the frequency of IS1 transposition is a function of glucose concentration in the growth medium, it is increased by streptomycin amounts that are below minimum inhibitory concentration (sub-MIC) and is inhibited in an rpsL150 strain with high translation accuracy. Since starved cells are known to enhance ribosome frameshifting, our data suggests that growth conditions applied in this study could affect IS1 transposition by increasing translation infidelity.

Protective Role for H-NS Protein in IS1 Transposition

ABSTRACT The transposase (InsAB′) of the insertion element IS1 can create breaks in DNA that lead to induction of the SOS response. We have used the SOS response to InsAB′ to screen for host mutations that affect InsAB′ function and thus point to host functions that contribute to the IS1 transposition mechanism. Mutations in the hns gene, which codes for a DNA binding protein with wide-ranging effects on gene expression, abolish the InsAB′-induced SOS response. They also reduce transposition, whether by simple insertion or cointegrate formation, at least 100-fold compared with the frequency seen in hns+ cells. Examination of protein profiles revealed that in an hns-null mutant, InsAB′ is undetectable under conditions where it constitutes the most abundant protein in hns+ cells. Likewise, brief labeling of the hns cells with [35S]methionine revealed very small amounts of InsAB′, and this was undetectable after a short chase. Transcription from the promoters used to express insAB′ was essentially unaltered in hns cells, as was the level of insAB′ mRNA. A mutation in lon, but not in ftsH or clpP, restored InsAB′ synthesis in the hns strain, and a mutation in ssrA partially restored it, implying that the absence of H-NS leads to a problem in completing translation of insAB′ mRNA and/or degradation of nascent InsAB′ protein.

Different reading frames are responsible for IS1-dependent deletions and recombination.

Two classes of plasmids in addition to the parent become apparent when plasmids that contain direct repeats of IS1. One class of plasmids has deleted sequences from the end of IS1 to nonrandom sites within the plasmid. The appearance of these plasmids in the population requires intact insA and insB reading frames, but not insC. The other class of plasmids has undergone an exchange within the direct repeats of IS1 on the plasmid. Their appearance requires InsC but neither InsA nor InsB. The two reactions may represent two distinguishable steps in IS1 transposition. The InsC-catalyzed exchange is independent of RecA and resembles hologous recombination since the frequency of recombinants arising from exchanges in different regions of IS1 appears to be roughly proportional to the size of the region. InsC can also catalyze an exchange between direct repeats of non-IS1 DNA.

MECHANISM OF IS1 TRANSPOSITION IN E. COLI: CHOICE BETWEEN SIMPLE INSERTION AND COINTEGRATION

ABSTRACT nsertion element IS 1 and IS 1-based transposon Tn 9 generate cointegrates (containing vector and target DNAs joined by duplicate copies of IS 1 or Tn 9) and simple insertions (containing IS 1 or Tn 9 detached from vector sequences). Based on studies of transposon Tn 5 we had proposed a conservative (non-replicative) model for simple insertion. Others had proposed that all transposition is replicative, occurring in a rolling circle structure, and that the way DNA strands are joined when replication terminates determines whether a simple insertion or a cointegrate is formed.—We selected for the transposition of amp and cam resistance markers from pBR322::Tn 9 plasmids to an F factor in recA - E. coli and identified products containing three and four copies of IS 1, corresponding to true cointegrates (from monomeric plasmids), and simple insertions (from dimeric plasmids). The simple insertions with four copies of IS 1 outnumbered those with three by a ratio of about 3:1, whereas true cointegrates containing three copies of IS 1 were more numerous than those with four.—A straightforward rolling circle model had predicted that the simple insertions containing three copies of IS 1 should be more frequent than those with four. Because we obtained the opposite result we propose that simple insertions only arise when the element fails to replicate or if replication starts but then terminates prematurely. The two classes of products, simple insertions and cointegrates, reflect alternative conservative and replicative fates, respectively, of an early intermediate in transposition.