The LacI-Type transcriptional regulator AraR acts as an L-arabinose-responsive repressor of L-arabinose utilization genes in Corynebacterium glutamicum ATCC 31831.
The Corynebacterium glutamicum ATCC 31831 araBDA operon consists of three l-arabinose catabolic genes, upstream of which the galM, araR, and araE genes are located in opposite orientation. araR encodes a LacI-type transcriptional regulator that negatively regulates the l-arabinose-inducible expression of araBDA and araE (encoding an l-arabinose transporter), through a mechanism that has yet to be identified. Here we show that the AraR protein binds in vitro to three sites: one upstream of araBDA and two upstream of araE. We verify that a 16-bp consensus palindromic sequence is essential for binding of AraR, using a series of mutations introduced upstream of araB in electrophoretic mobility shift assays. Moreover, the DNA-binding activity of AraR is reduced by l-arabinose. We employ quantitative reverse transcription-PCR (qRT-PCR) analyses using various mutant strains deficient in l-arabinose utilization genes to demonstrate that the prominent upregulation of araBDA and araE within 5 min of l-arabinose supplementation is dependent on the uptake but independent of the catabolism of l-arabinose. Similar expression patterns, together with the upregulation by araR disruption without l-arabinose, are evident with the apparent galM-araR operon, although attendant changes in expression levels are much smaller than those realized with the expression of araBDA and araE. The AraR-binding site upstream of araB overlaps the -10 region of the divergent galM promoter. These observations indicate that AraR acts as a transcriptional repressor of araBDA, araE, and galM-araR and that l-arabinose acts as an intracellular negative effector of the AraR-dependent regulation.
Research Institute of Innovative Technology for the Earth, Kizugawadai, Kizugawa, Kyoto, Japan Graduate School of Biological Sciences, Nara Institute of Science and Technology, Takayama, Ikoma, Nara, Japan.
Takayuki
T
Kuge
Research Institute of Innovative Technology for the Earth, Kizugawadai, Kizugawa, Kyoto, Japan.
Haruhiko
H
Teramoto
Research Institute of Innovative Technology for the Earth, Kizugawadai, Kizugawa, Kyoto, Japan.
Hideaki
H
Yukawa
Research Institute of Innovative Technology for the Earth, Kizugawadai, Kizugawa, Kyoto, Japan Graduate School of Biological Sciences, Nara Institute of Science and Technology, Takayama, Ikoma, Nara, Japan mmg-lab@rite.or.jp.
Masayuki
M
Inui
D001089:Arabinose / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D048230:Corynebacterium glutamicum / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D056968:Lac Repressors / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding
D001426:Bacterial Proteins; D056968:Lac Repressors; D001089:Arabinose