The laci-type transcriptional regulator AraR acts as an L-arabinose-responsive repressor of L-arabinose utilization genes in Corynebacterium glutamicum ATCC 31831

Abstract

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. © 2014, American Society for Microbiology.