Hopping into a hot seat: Role of DNA structural features on IS5-mediated gene activation and inactivation under stress

Abstract

Insertion sequence elements (IS elements) are proposed to play major roles in shaping the genetic and phenotypic landscapes of prokaryotic cells. Recent evidence has raised the possibility that environmental stress conditions increase IS hopping into new sites, and often such hopping has the phenotypic effect of relieving the stress. Although stress-induced targeted mutations have been reported for a number of E. coli genes, the glpFK (glycerol utilization) and the cryptic bglGFB (β-glucoside utilization) systems are among the best characterized where the effects of IS insertion-mediated gene activation are well-characterized at the molecular level. In the glpFK system, starvation of cells incapable of utilizing glycerol leads to an IS5 insertion event that activates the glpFK operon, and enables glycerol utilization. In the case of the cryptic bglGFB operon, insertion of IS5 (and other IS elements) into a specific region in the bglG upstream sequence has the effect of activating the operon in both growing cells, and in starving cells. However, a major unanswered question in the glpFK system, the bgl system, as well as other examples, has been why the insertion events are promoted at specific locations, and how the specific stress condition (glycerol starvation for example) can be mechanistically linked to enhanced insertion at a specific locus. In this paper, we show that a specific DNA structural feature (superhelical stress-induced duplex destabilization, SIDD) is associated with “stress-induced” IS5 insertion in the glpFK, bglGFB, flhDC, fucAO and nfsB systems. We propose a speculative mechanistic model that links specific environmental conditions to the unmasking of an insertional hotspot in the glpFK system. We demonstrate that experimentally altering the predicted stability of a SIDD element in the nfsB gene significantly impacts IS5 insertion at its hotspot.

Insertion Sequence (IS) Element-Mediated Activating Mutations of the Cryptic Aromatic β-Glucoside Utilization (BglGFB) Operon Are Promoted by the Anti-Terminator Protein (BglG) in Escherichia coli

“…Several non-canonical DNA structures have been identified such as melted bubbles, Z-DNA, H-DNA and G quadruplexes in both prokaryotic and eukaryotic genomes [43][44][45][46][47]. One major representative of DNA structures is the SIDD (superhelical stress-induced DNA duplex destabilization) structures that are preferentially present in regulatory regions [48,49] and specifically promote insertion of IS elements [16]. A strong SIDD structure is present upstream of the bgl promoter [16], and the majority of IS insertional mutations occur in this SIDD region [22,50,51].…”

“…It appears that such examples are not uncommon in both bacteria and eukaryotes [13]. Additional examples in bacteria include IS5 activation of the fucose AO (fucAO) operon in E. coli [14], furazolidone sensitivity A and B (nfsA and nfsB) inactivation by IS elements, also in E. coli [15,16], the regulation of capsule biosynthesis in Neisseria meningitidis [17], developmentally programmed transposon excision in the Bacillus subtilis sporulation cascade [12], drug (piperacillin/tazobactam) resistance in E. coli [18], and retrotransposon-regulated gene expression in eukaryotes [19,20]. Thus, there is an ever-increasing number of examples of the regulation of gene expression and function that may be subject to transposon-mediated directed or adaptive mutation.…”

Bioassay for Determining the Concentrations of Caffeine and Individual Methylxanthines in Complex Samples

“…This pDCAF3-IS5 plasmid variant still maintains sufficient demethylation activity to support caffeine measurement by ⌬guaB E. coli in the manner described by Quandt et al (22). The IS5 insertion presumably alters expression from the interrupted promoter, as has been observed in other studies (30), potentially making this plasmid less burdensome than the original pDCAF3 plasmid.…”

Evolution of Bacterial Cross-Resistance to Lytic Phages and Albicidin Antibiotic

“…It has been previously assumed that insertion and excision of IS elements are stochastic events. However, recent evidence suggests that IS movement may actually be a form of directed mutagenesis in which IS elements target specific chromosomal loci to reduce stress under certain environmental conditions (Humayun et al, 2017). The isolation of phage-resistant mutants in this study relied on spontaneous mutations accumulated in an overnight culture, while the isolation of antibiotic-resistant mutants allowed for both selection of spontaneous resistant mutants as well as evolution in the presence of albicidin.…”