Expression of the arsenite oxidation regulatory operon in Rhizobium sp. str. NT‐26 is under the control of two promoters that respond to different environmental cues

Abstract

Rhizobium sp. str. NT‐26 is a Gram‐negative facultative chemolithoautotrophic arsenite oxidizer that has been used as a model organism to study various aspects of arsenite oxidation including the regulation of arsenite oxidation. The three regulatory genes, aioX, aioS, and aioR, are cotranscribed when NT‐26 was grown in the presence or absence of arsenite. The aioXSR operon is upregulated in stationary phase but not by the presence of arsenite in the growth medium. The two transcription start sites upstream of aioX were determined which led to the identification of two promoters, the housekeeping promoter RpoD and the growth‐phase‐dependent promoter RpoE2. Promoter–lacZ fusions confirmed their constitutive and stationary phase expressions. The involvement of the NT‐26 sigma factor RpoE2 in acting on the NT‐26 RpoE2 promoter was confirmed in vivo in Escherichia coli, which lacks a rpoE2 homolog, using a strain carrying both the promoter–lacZ fusion and the NT‐26 rpoE2 gene. An in silico approach was used to search for other RpoE2 promoters and AioR‐binding motifs and led to the identification of other genes that could be regulated by these proteins including those involved in quorum sensing, chemotaxis, and motility expanding the signaling networks important for the microbial metabolism of arsenite.

Transcriptomics analysis defines global cellular response ofAgrobacterium tumefaciens5A to arsenite exposure regulated through the histidine kinases PhoR and AioS

“…Of the 148 genes influenced in the Δ aioS mutant, only slightly over half (78 genes) were associated with one or the other putative AioR‐type binding sites (Supporting Information Table 2). The Corsini and colleagues () motif was associated with only 18 of these genes, whereas the motif described by Shi and colleagues () was found associated with 67 genes; seven genes were found to be associated with both motifs. As would be predicted, the degenerate sequence of Shi and colleagues (), allowed for greater flexibility and thus it was identified more frequently.…”

“…The predicted and experimentally demonstrated degenerate AioR binding site motif (Li et al ., ; Shi et al ., ) and that predicted by Corsini and colleagues () were used to search for potential AioR binding sites (see Supporting Information Table 2). Of the 148 genes influenced in the Δ aioS mutant, only slightly over half (78 genes) were associated with one or the other putative AioR‐type binding sites (Supporting Information Table 2).…”

“…It was of interest to determine if the genes influenced by AioS might also be physically linked with σ 54 and AioR binding sites, which are viewed to be essential for the expression of the aioBA genes (encode As III oxidase) (Kang et al ., ; Corsini et al ., ; Shi et al ., ). As expected, both were found upstream of aioB (Supporting Information Table 2), but this was not the case for most of the genes perturbed in the aioS mutant (Supporting Information Table 2).…”

“…FIMO utilizes log‐posterior odds scoring and position‐specific priors to search for a given motif (Grant et al ., ; Cuellar‐Partida et al ., ). The well‐defined σ 54 consensus sequence, TGG(N9)TGC, (Cannon et al ., ; Kang et al ., ) and proposed AioR binding sequences GTCCGCAAAATCAGGACA (Corsini et al ., ) and GT[TC][AC][CG][GCT][AG][AG]A[ACT][CGA][GCT][GTA]AAC (Shi et al ., ) were used as search inputs. Only motifs with a p value ≤0.001 were considered as putative binding sites.…”

“…An important assumption going into this analysis was that transcriptional control of all aioSinfluenced genes would also involve AioR, the cognate regulatory protein for AioS. Targeted motif searches included binding sites for AioR (Corsini et al, 2017;Shi et al, 2018) and σ 54 , which is known to be involved in transcriptional regulation of arsenite oxidase in 5A and Herminiimonas arsenicoxydans Kang et al, 2012a). Searches were conducted within the 300 nucleotide region upstream of the translational start site for each gene.…”