Positive Feedback Regulation of stgR Expression for Secondary Metabolism in Streptomyces coelicolor

Abstract

LysR-type transcriptional regulators (LTTRs) compose a large family and are responsible for various physiological functions in bacteria, while little is understood about their regulatory mechanism on secondary metabolism in Streptomyces. Here we reported that StgR, a typical LTTR in Streptomyces coelicolor, was a negative regulator of undecylprodigiosin (Red) and ␥-actinorhodin (Act) production in the early developmental phase of secondary metabolism by suppressing the expression of two pathway-specific regulator genes, redD and actII-orf4, respectively. Meanwhile, stgR expression was downregulated during secondary metabolism to remove its repressive effects on antibiotic production. Moreover, stgR expression was positively autoregulated by direct binding of StgR to its own promoter (stgRp), and the binding site adjacent to translation start codon was determined by a DNase I footprinting assay. Furthermore, the StgR-stgRp interaction could be destroyed by the antibiotic ␥-actinorhodin produced from S. coelicolor. Thus, our results suggested a positive feedback regulatory mechanism of stgR expression and antibiotic production for the rapid and irreversible development of secondary metabolism in Streptomyces. LysR-type transcriptional regulators (LTTRs), first named after LysR, a transcriptional activator of lysA in Escherichia coli (1), have expanded considerably to the largest family after 3 decades and spread ubiquitously in bacteria. They have been structurally well characterized to have a conserved helix-turn-helix (HTH) motif at the N terminus for DNA binding and a regulatory domain for substrate or inducer binding at the C terminus (2). Consistent with their wide distributions and great quantities, they have diverse and conserved regulatory functions in bacteria for primary metabolism (3, 4), secondary metabolism (5, 6), stress responses (7), cell division (8), virulence (9, 10), protection (11), etc. Typically, most LTTRs function as the global transcriptional regulators by directly binding to the promoters of their regulons. Upon signaling, the assimilated extracellular ligands or produced intracellular metabolites can act as substrates or inducers to interact with the C-terminal domain to cause conformation changes to influence the DNA-binding affinity of LTTRs (2, 12).Species of Streptomyces, the soil-dwelling Gram-positive bacteria, are well-known for their complex morphogenesis and secondary metabolism. Among their abundant secondary metabolites, antibiotics are produced with patent clinical or commercial applications (13). The onset of secondary metabolism is triggered by environmental stimuli, and subsequent intracellular signaling pathways are equally required for felicitous development of secondary metabolism (14). Nutrients, including N-acetylglucosamine, and autoregulatory factors, such as ␥-butyrolactones, can regulate antibiotic production through DasR-and AdpA-mediated signaling pathways, respectively (14, 15). Other global regulatory systems, including the two-component system (TCS), ppGpp, ...

Discovery of scmR as a global regulator of secondary metabolism and virulence in Burkholderia thailandensis E264

“…Streptomyces coelicolor contains an abundance of LTTRs and one of these, StgR, has been shown to limit production of actinorhodin and prodigiosin in the early growth phase via interaction with pathwayspecific transcriptional regulators (44). In stationary phase cultures, when biosynthesis of these metabolites is activated, stgR expression was found to be down-regulated.…”

Transcriptional Regulation of the Daptomycin Gene Cluster in Streptomyces roseosporus by an Autoregulator, AtrA

“…These probes were gel-purified and eluted in sterile water. EMSA was demonstrated as described previously with 1 ng of probes and an increasing amount of protein (24).…”

“…DNase I Footprinting Assay-The DNase I footprinting assays were demonstrated as described previously (24). 5Ј-FAMlabeled dptEp and atrAp were prepared by PCR from plasmids pTA2-dptEp and pTA2-atrAp, respectively, with universal primers 29 ϩ 23 and then gel-purified.…”

DepR1, a TetR Family Transcriptional Regulator, Positively Regulates Daptomycin Production in an Industrial Producer, Streptomyces roseosporus SW0702

“…The DNase I footprinting assays were performed as previously described (29). Briefly, 5=-6-carboxyfluorescein (FAM)-labeled dptEp and depR1p were prepared by PCR from pUC18-dptEp and pUC18-depR1p with primers 20 and 21.…”

“…Electrophoretic mobility shift assays (EMSAs) were performed as described previously (29). The 5=-biotin-labeled probes were obtained by PCR from pUC18, pUC18-dptEp, and pUC18-depR1p using primers 19 and 20.…”