The LacI-Type Transcriptional Regulator AraR Acts as an l -Arabinose-Responsive Repressor of l -Arabinose Utilization Genes in Corynebacterium glutamicum ATCC 31831

Abstract

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.