Transcriptional regulation of the iac locus from Acinetobacter baumannii by the phytohormone indole-3-acetic acid

Abstract

The iac locus is involved in indole-3-acetic acid (IAA) catabolism in Acinetobacter baumannii. Nine structural genes of iac are transcribed in the same direction, whereas iacR, which encodes a MarR-type transcriptional regulator, is transcribed in the opposite direction. The IacA protein, which is encoded by the second structural gene of the iac locus, is expressed in an IAA-dependent manner. Here, we characterized gene expression from this locus in wild type A. baumannii and in an iacR mutant; this revealed that the iacH promoter is negatively regulated by IacR. The transcriptional site of iacH was determined by using 5' rapid amplification of cDNA ends; one IacR-binding site was identified between positions -35 and +28 of the iacH promoter. Sequence analysis and an electrophoretic mobility shift assay indicated that recombinant IacR binds specifically to a sequence with dyad symmetry in the iacR-iacH overlapping promoters in the absence of IAA. In addition, a two-plasmid expression system in Escherichia coli showed that IAA probably serves as a ligand that binds to IacR and releases it from the iacH promoter, thereby allowing RNA polymerase to transcribe iac. Thus, iac is expressed in order to promote IAA degradation, whereas free IacR is required for iac repression. We conclude that IacR serves as a key regulator of IAA degradation in A. baumannii in the rhizosphere. These results provide new insights into the possible role of A. baumannii in the environment.

Biochemical and Genetic Bases of Indole-3-Acetic Acid (Auxin Phytohormone) Degradation by the Plant-Growth-Promoting Rhizobacterium Paraburkholderia phytofirmans PsJN

“…Comparisons between iac homologues showed amino acid identities ranging from 38 to 62% (see Table S1 in the supplemental material). Only the iacR gene strain 1290 homologue, a transcriptional repressor belonging to the MarR family (14,15), is absent in the strain PsJN genome. The strain PsJN iac gene sequences are adjacent to other genes, not found in P. putida 1290, possibly playing regulatory and transport roles in IAA degradation (Fig.…”

“…IAA degradation by strain PsJN apparently uses some functionally redundant genes and other genes not previously reported in P. putida 1290, encoding transport and tautomerase superfamily enzymes. We also reported that, in contrast with P. putida and A. baumannii, where iac gene regulation is controlled by a MarR-type transcriptional repressor responding to the IAA molecule (14,15), IAA degradation in P. phytofirmans is regulated by a two-component system and a LysR-type activator, which apparently respond to IAA in combination with DOAA, the previously proposed intermediate in strain 1290 (14).…”

“…These primers contain a 20-bp terminal sequence homologous to the terminus of the fragment to be linked, and the sequences were combined and ligated to generate a new DNA molecule in a one-step isothermal reaction (66). Also, PCR products lacking one or two of these iac genes were amplified using primer pairs listed in Table 2: iacBIHE (primer pair 2-11), iacCD (primer pair 3-4), iacG (primer pair 5-6), and iacF (primer pair 7-8) genes were used to obtain the iac1ΔA derivative; iacA (primer pair 1-12), iacIHE (primer pair 2-13), iacCD (primer pair 3-4), iacG (primer pair 5-6), and iacF (primer pair 7-8) genes were used to produce the iac1ΔB derivative; iacABIHE (primer pair 1-15), iacG (primer pair 5-6), and iacF (primer pair 7-8) genes were used to produce the iac1ΔCD derivative; iacABIH (primer pair [1][2][3][4][5][6][7][8][9][10][11][12][13][14], iacCD (primer pair 3-4), iacG (primer pair 5-6), and iacF (primer pair 7-8) genes were used to give the iac1ΔE derivative; iacABIHE (primer pair 1-2), iacCD (primer pair 3-4), and iacG (primer pair 5-16) genes were used to create the iac1ΔF derivative; iacABIHE (primer pair 1-2), iacCD (primer pair 3-4), and iacF (primer pair 8-17) genes were used to generate the iac1ΔG derivative; iacABI (primer pair [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18], iacE (primer pair [2][3][4]…”

“…Compared with P. putida 1290, where the iac gene cluster includes only a transcriptional repressor belonging to the MarR family, named iacR (14,15), which is absent in the genome of P. phytofirmans PsJN, the regulation of IAA catabolism in strain PsJN is far more complex: a histidine kinase signal transducer and a LuxR family protein receptor (a putative two-component system), iacS and iacR1, respectively, and a LysR-type transcriptional regulator (LTTR), iacR2 (Fig. 1B).…”

“…1A) (12,14). More recently, the iacR gene from Acinetobacter baumannii, encoding a putative MarR-type transcriptional regulator, was reported to control iac gene repression in the absence of IAA (15). The roles of other iac genes remain unknown.…”