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

Abstract

The glutamine synthetase II (GSII, encoded by glnII) activity detectable in crude extracts from Streptomyces coelicolor is low compared to the activity of glutamine synthetase I (GSI, encoded by glnA) and to that of GSII from S. viridochromogenes. We have identified and sequenced a 3.9-kb BglII-BamHI fragment carrying the glutamine synthetase II gene (glnII) from S. coelicolor. Besides glnII, this region contains four ORFs (orf1-orf4). While homologues of orf1 and orf2 were also found in the glnII region of the S. viridochromogenes chromosome, this was not the case for orf3 and orf4, which encode 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 first comprises a vegetative promoter sequence and the second contains sequence elements that are recognized by Esigma31. Similar promoter structures were found upstream of the S. viridochromogenes glnII gene. The involvement of ptr in glnII regulation was studied by gel retardation assays. Recombinant Ptr interacted with the upstream region of ptr, but not with the promoter region of glnII. A ptr gene replacement mutant (S. coelicolor IP) was also constructed. RT-PCR analysis of RNA from wild-type S. coelicolor and the IP mutant demonstrated that expression of orf3 depends on Ptr. Thus, the difference in gene organization between S. coelicolor and S. viridochromogenes is not responsible for the difference in GSII activity.

Phosphate control over nitrogen metabolism in Streptomyces coelicolor: direct and indirect negative control of glnR, glnA, glnII and amtB expression by the response regulator PhoP

“…The main promoter should correspond to the major vegetative σ factor and its transcription start site is the adenine −79. The minor promoter shows putative elements of σ 31 (14). This sigma factor, which is more active at stationary phase, recognizes promoters of actIII (an actinorhodin structural gene) and of hrdD (a homologue of Escherichia coli rpoD ), and also the glnR P2 promoter (31).…”

“…The overall nitrogen metabolism is regulated in S. coelicolor by complex mechanisms that involve an apparent doubling of some structural and regulatory genes (12). There are two glutamine synthetase (GS) genes in S. coelicolor : glnA (13), that encodes the GS type I (GS-I), orthologous to that found in prokaryotes, and glnII (14), which encodes GS type II (GS-II), similar to eukaryotic GS. The glnII gene appears to be present in all Streptomyces species (15), but it is absent in the related actinomycetes Mycobacterium and Corynebacterium (12).…”

Gamma-Glutamylpolyamine Synthetase GlnA3 Is Involved in the First Step of Polyamine Degradation Pathway in Streptomyces coelicolor M145

“…Consequently, some proteins annotated as glutamine synthetase-like enzymes (GS-like) may represent still not recognized γ-glutamylpolyamine synthetases. Indeed, Streptomyces coelicolor A3(2) harbors two genes, glnA ( SCO2198 ) and glnII ( SCO2210 ), encoding GSI and GSII, whose function and regulation was extensively studied (Hillemann et al, 1993; Fink et al, 1999; Weisschuh et al, 2000) as well as three other genes, glnA2 ( SCO2241 ), glnA3 ( SCO6962 ), and glnA4 ( SCO1613 ), annotated as encoding putative GS-like enzymes (Rexer et al, 2006). In silico analysis of all glnA -like genes: glnA2 , glnA3 , and glnA4 across the actinobacterial genomes revealed that glnA has evolved to specialized glnA -like genes encoding proteins that might be involved in the colonization and survival in many diverse habitats (Hayward et al, 2009).…”

Nitrogen Metabolism in <i>Streptomyces coelicolor</i>: Transcriptional and Post-Translational Regulation

“…The glnA gene is transcribed at a constant level throughout development and GSI activity can be detected in all growth phases . The glnII gene encodes the GSII which shows high sequence similarity to eukaryotic-type GS enzymes Weisschuh et al, 2000] . GSII enzymes are characterized as heat-labile homooctamers and are typical for eukaryotes.…”