Abstract
Transposable Insertion Sequences (IS elements) have been shown to provide various benefits to their hosts via gene activation or inactivation under stress conditions by appropriately inserting into specific chromosomal sites. Activation is usually due to derepression or introduction of a complete or partial promoter located within the element. Here we define a novel mechanism of gene activation by the transposon IS5 in Escherichia coli. The glycerol utilization operon, glpFK, that is silent in the absence of the cAMP-Crp complex, is activated by IS5 when inserted upstream of its promoter. High-level expression is nearly constitutive, only mildly dependent on glycerol, glucose, GlpR, and Crp, and allows growth at a rate similar to or more rapid than that of wild-type cells. Expression is from the glpFK promoter and dependent on (1) the DNA phase, (2) integration host factor (IHF), and (3) a short region at the 3′ end of IS5 harboring a permanent bend and an IHF binding site. The lacZYA operon is also subject to such activation in the absence of Crp. Thus, we have defined a novel mechanism of gene activation involving transposon insertion that may be generally applicable to many organisms.
Highlights
Living organisms possess a variety of mutagenic means to generate genetic diversity, and these depend on environmental conditions and genomic composition [1,2]
We demonstrate for the first time that one such transposon, Insertion Sequence 5 (IS5), when positioned upstream of a metabolic operon of E. coli, can activate the otherwise cryptic expression of the operon
This effect is due solely to a short region at the 39 end of IS5 that harbors a permanent bend and an overlapping nucleoid protein binding site, both of which are required for maximal gene expression
Summary
Living organisms possess a variety of mutagenic means to generate genetic diversity, and these depend on environmental conditions and genomic composition [1,2]. One transposon-mediated mechanism of gene activation involves the formation of a ‘‘hybrid promoter’’ when a small transposon, an Insertion Sequence (IS) element, inserts into the promoter region. In this case, insertion of an IS element results in placing an outwardly directed 235 hexamer in one of the terminal inverted repeats (IRs) of the transposon at the correct distance from a resident 210 hexamer. Activation can occur by initiating transcription within the transposon, traversing the terminal IR and reading the gene of interest This second type of mechanism has been observed for IS3 [6] and IS10 [7]
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