Genetic Evidence that GTP Is Required for Transposition of IS 903 and Tn 552 in Escherichia coli

Abstract

Surprisingly little is known about the role of host factors in regulating transposition, despite the potentially deleterious rearrangements caused by the movement of transposons. An extensive mutant screen was therefore conducted to identify Escherichia coli host factors that regulate transposition. An E. coli mutant library was screened using a papillation assay that allows detection of IS903 transposition events by the formation of blue papillae on a colony. Several host mutants were identified that exhibited a unique papillation pattern: a predominant ring of papillae just inside the edge of the colony, implying that transposition was triggered within these cells based on their spatial location within the colony. These mutants were found to be in pur genes, whose products are involved in the purine biosynthetic pathway. The transposition ring phenotype was also observed with Tn552, but not Tn10, establishing that this was not unique to IS903 and that it was not an artifact of the assay. Further genetic analyses of purine biosynthetic mutants indicated that the ring of transposition was consistent with a GTP requirement for IS903 and Tn552 transposition. Together, our observations suggest that transposition occurs during late stages of colony growth and that transposition occurs inside the colony edge in response to both a gradient of exogenous purines across the colony and the developmental stage of the cells.Transposons are defined as distinct regions of DNA that have the ability to move from one genomic location to another, a process mediated by element-encoded transposases (8, 9). While the main focus of transposition research has been in determining the detailed mechanisms of transposition, relatively little attention has been paid to how transposition is regulated in vivo or the in vivo requirements for the process (35). Transposon movement can result in gene inactivation (by insertional inactivation or deletion) or activation (by creation or introduction of promoters upstream of genes). In addition, intramolecular transposition can result in extensive DNA rearrangements, including deletions, inversions, and duplications of chromosome segments; these events are thought to play an important role in facilitating genome evolution (7-9). As the consequences of transposition can be either dire or favorable for the host organism, it is intuitive that the host has the capability to regulate this process. There are multiple examples of how transposons regulate their own movement, but very little work has focused on the role played by the host in regulating transposition. To address this, we have generated an insertion mutant library of Escherichia coli, which we have used to screen for host genes involved in regulation of IS903 transposition (E. Twiss, A. Coros, N. Tavakoli, and K. M. Derbyshire, unpublished data).The first comprehensive genetic screen for host mutants affecting transposition was carried out in 1985; it revealed a role for dam methylation in decreasing the rate of IS10, IS903, and IS50 transposi...

Transposition is modulated by a diverse set of host factors in Escherichia coli and is stimulated by nutritional stress

“…These, and previous, studies have also indicated that transposition of IS903 occurs at late times during colony growth (Fig. 2, and Coros et al, 2005).…”

“…Mutations received, or created by λRed mutagenesis were transduced into DH10B or HT321. For transduction of mutations into the recA – strain, DH10B, the recA gene was supplied in trans from a plasmid, pET001 (Coros et al ., 2005). Replication of this plasmid is temperature‐sensitive.…”

σ B Regulates IS 256 -Mediated Staphylococcus aureus Biofilm Phenotypic Variation

“…In particular, stress situations such as carbon starvation, extreme temperature, and UV light have been correlated with the enhanced transposition of bacterial mobile elements (10,15,22,31,33,39). These findings support the idea that bacteria derepress the movement of mobile elements as an adaptive response to stress and to permit new traits to evolve (6,46).…”

Transposons Tn 10 and Tn 5

“…This is an aspect of bacterial transposition that has not been thoroughly studied even though there are many (mostly anecdotal) examples of transposition systems responding to stress conditions. One exception is the IS903 system, in which the response of this element to defects in purine metabolism has been worked out reasonably well (28). Outside of host regulation there has been some new activity directed at further defining molecular details of the Tn5 and Tn10 hairpin reactions.…”