The RNA Polymerase Omega Factor RpoZ Is Regulated by PhoP and Has an Important Role in Antibiotic Biosynthesis and Morphological Differentiation in Streptomyces coelicolor

Abstract

The RNA polymerase (RNAP) omega factor () forms a complex with the ␣ 2 ␤␤ core of this enzyme in bacteria. We have characterized the rpoZ gene of Streptomyces coelicolor, which encodes a small protein (90 amino acids) identified as the omega factor. Deletion of the rpoZ gene resulted in strains with a slightly reduced growth rate, although they were still able to sporulate. The biosynthesis of actinorhodin and, particularly, that of undecylprodigiosin were drastically reduced in the ⌬rpoZ strain, suggesting that expression of these secondary metabolite biosynthetic genes is dependent upon the presence of RpoZ in the RNAP complex. Complementation of the ⌬rpoZ mutant with the wild-type rpoZ allele restored both phenotype and antibiotic production. Interestingly, the rpoZ gene contains a PHO box in its promoter region. DNA binding assays showed that the phosphate response regulator PhoP binds to such a region. Since luciferase reporter studies showed that rpoZ promoter activity was increased in a ⌬phoP background, it can be concluded that rpoZ is controlled negatively by PhoP, thus connecting phosphate depletion regulation with antibiotic production and morphological differentiation in Streptomyces.In bacteria, the RNA polymerase (RNAP) complex plays a central role in transcription and is a target for regulation of primary metabolism (6,7,44). The rpoZ gene encodes the RNAP omega () subunit, which forms a complex with the ␣ 2 ␤␤Ј core of this enzyme. The subunit has been identified in the RNAPs of most free-living bacteria. This protein is functionally homologous to the RpoK subunit of the archaeal RNA polymerase complex and the RPB6 subunit of the eukaryotic RNA polymerases I, II, and III (32). In Escherichia coli the subunit interacts with the ␤Ј subunit and promotes assembly of the RNA polymerase complex (14, 32), although it is not essential for survival in this bacterium (13).Streptomyces spp. are soil-dwelling bacteria that are notorious for their ability to produce thousands of antibiotics, pigments, antitumor agents, immunomodulators, and a variety of other bioactive secondary metabolites (1, 2, 8). Differential expression of secondary metabolism genes occurs following nutrient depletion (34), but the transcriptional control mechanisms that govern the onset of secondary metabolites are still obscure (29). The rpoZ gene of Streptomyces kasugaensis has been shown to be required for antibiotic production and morphological differentiation but is not essential for growth (21). A DNA fragment containing the rpoZ gene was shown to complement an S. kasugaensis pleiotropic mutant deficient in aerial mycelium formation and kasugamycin biosynthesis. Although sigma factors in Streptomyces have received considerable attention in relation to the expression of antibiotic biosynthetic genes (9,19,20), the role of the RNAP subunit is still obscure.The expression of many genes involved in antibiotic biosynthesis is negatively controlled by the phosphate concentration in the medium (reviewed in references 28 and 30). Limitati...

Molecular Regulation of Antibiotic Biosynthesis in Streptomyces

“…subunit of RNA polymerase, yet RpoZ is required for normal levels of production of ACT and RED (72). It is notable that the effects of phoP deletion on Streptomyces lividans, a very close relative of S. coelicolor, were quite different: in that organism, in which ACT and RED production is turned off under most conditions, phoP deletion caused strongly increased production of the two antibiotics (73).…”

“…The two-component system PhoR-PhoP (which is widely distributed across prokaryotes) is the major signal transduction system for phosphate control in S. coelicolor and affects ACT and RED production by (directly or indirectly) influencing the transcription of act and red genes (72,73). Similar effects on antibiotic production in other streptomycetes, including S. griseus (candicidin) (74,75), Streptomyces natalensis (pimaricin) (76), and Streptomyces rimosus (oxytetracycline) (75,77), imply that this role of the PhoR-PhoP system is widespread.…”

The master regulator PhoP coordinates phosphate and nitrogen metabolism, respiration, cell differentiation and antibiotic biosynthesis: comparison in Streptomyces coelicolor and Streptomyces avermitilis

“…This finding indicates that binding of PhoP to the promoter region prevents reading by the RNA polymerase. 21 The afsS gene, which is well known to be recognized by PhoP in S. coelicolor, affects avermectin biosynthesis in S. avermitilis 8 and this suggests that one of the indirect targets of PhoP that controls avermectin biosynthesis is the PhoP regulation of afsS. Indeed, the new bioinformatic analysis (Table 2) reveals that there is a well conserved PHO box in the S. avermitilis afsS promoter that overlaps with the AfsR binding site.…”

“…In fact, when PhoP binds to the − 35 region of the promoters of PhoP-regulated genes, it acts as an activator by recruiting RNA polymerase to the promoter regions, whereas when it binds to the − 10 regions, or inside the open reading frame, usually it works as a repressor by preventing transcription. [19][20][21] THE pho REGULON: PhoP-REGULATED GENES Transcriptomic studies, footprinting assays and in vivo expression studies with reporter genes allowed the initial identification of about 50 genes, which are regulated by PhoP and form the core of the pho regulon. 17 A list of relevant genes regulated by PhoP is shown in Table 1 and a summary of downregulated and upregulated genes is shown in Figure 2.…”

The ω Subunit Governs RNA Polymerase Stability and Transcriptional Specificity in Staphylococcus aureus

“…Indeed, none of these studies have unraveled the molecular basis for alterations in mutant strains, meaning that the role of in Gram-positive species is still relatively elusive. Those effects that have been detailed for rpoZ mutants include alterations in cell wall morphology, cell motility, protein secretion, and biofilm formation (16,(26)(27)(28)(29). Importantly, the role of this subunit in the virulence of pathogenic species has yet to be evaluated.…”