Transcriptional analysis of the gene for glutamine synthetase II and two upstream genes in Streptomyces coelicolor A3(2)

The glutamine synthetase IT (GSII, encoded by glnII) activity detectable in crude extracts from Streptomyces coelicolor is low compared to the activit y of glutamine synthetase I (GSI, encoded by glnA) and to that of GSII fi o m S. viridochromogenes. We have identified and sequenced a 3.9-kb Bg/II-Bam HI fragment carrying the glutamine synthetase II gene (glnII) from S. coeli color. Besides glnII, this region contains four ORFs (orf1-orf4). While hom ologues of orf1 and orf2 were also found in the glnII region of the S. viri dochromogenes chromosome, this was not the case for orf3 and orf4, which en code a putative hydrolase and a transcriptional regulator (Ptr) of the MarR family, respectively. High-resolution S1 nuclease mapping showed that the S. coelicolor glnII gene is expressed from two overlapping promoters. The f irst comprises a vegetative promoter sequence and the second contains seque nce elements that are recognized by E sigma (31). Similar promoter structur es were found upstream of the S. viridochromogenes glnII gene. The involvem ent of ptr in glnII regulation was studied by gel retardation assays. Recom binant Ptr interacted with the upstream region of ptr.. but not with the pr omoter region of glnII. A ptr gene replacement mutant (S. coelicolor IF) wa s also constructed. RT-PCR analysis of RNA from wild-type S, coelicolor and the IP mutant demonstrated that 0expression of orf3 depends on Ptr. Thus, the difference in gene organization between S, coelicolor and S. viridochro mogenes is not responsible for the difference in GSII activity.