Activation of Dormant Bacterial Genes byNonomuraeasp. Strain ATCC 39727 Mutant-Type RNA Polymerase

Abstract

There is accumulating evidence that the ability of actinomycetes to produce antibiotics and other bioactive secondary metabolites has been underestimated due to the presence of cryptic gene clusters. The activation of dormant genes is therefore one of the most important areas of experimental research for the discovery of drugs in these organisms. The recent observation that several actinomycetes possess two RNA polymerase ␤-chain genes (rpoB) has opened up the possibility, explored in this study, of developing a new strategy to activate dormant gene expression in bacteria. Two rpoB paralogs, rpoB(S) and rpoB(R), provide Nonomuraea sp. strain ATCC 39727 with two functionally distinct and developmentally regulated RNA polymerases. The product of rpoB(R), the expression of which increases after transition to stationary phase, is characterized by five amino acid substitutions located within or close to the so-called rifampin resistance clusters that play a key role in fundamental activities of RNA polymerase. Here, we report that rpoB(R) markedly activated antibiotic biosynthesis in the wild-type Streptomyces lividans strain 1326 and also in strain KO-421, a relaxed (rel) mutant unable to produce ppGpp. Site-directed mutagenesis demonstrated that the rpoB(R)-specific missense H426N mutation was essential for the activation of secondary metabolism. Our observations also indicated that mutant-type or duplicated, rpoB often exists in nature among rare actinomycetes and will thus provide a basis for further basic and applied research.

Rare earth elements activate the secondary metabolite–biosynthetic gene clusters in Streptomyces coelicolor A3(2)

“…The mechanism of action, however, remains to be clarified. The present method, together with other methods reported recently, 8,9,12,[22][23][24] may be useful for activating silent genes, eventually leading to the discovery of novel biologically active compounds. Figure 3 Comparative metabolic profiling of the culture extracts.…”

Activation and Products of the Cryptic Secondary Metabolite Biosynthetic Gene Clusters by Rifampin Resistance ( rpoB ) Mutations in Actinomycetes

“…The mutation at position H437 has been shown to circumvent the detrimental effects of the relA and afsB mutations (in S. coelicolor) and the relC mutation (in S. lividans) on actinorhodin production, perhaps by mimicking the ppGpp-bound form of RNAP (40)(41)(42)(43). Taken together, ppGpp likely participates significantly even in the activation of cryptic gene clusters as well as highly expressed, well-known, secondary metabolite gene clusters.…”

Upregulation of the Phthiocerol Dimycocerosate Biosynthetic Pathway by Rifampin-Resistant,rpoBMutant Mycobacterium tuberculosis

“…However, relatively little is known about M. tuberculosis adaptive mechanisms, which may compensate for mutations in rpoB. Recently, compensatory mutations in RNA polymerase genes in rpoB mutant isolates of M. tuberculosis have been identified by comparative genomics (18).Specific gene upregulation associated with rpoB mutation has been observed in numerous model organisms, including Streptomyces spp., which are environmental organisms phylogenetically related to M. tuberculosis (35,37,69). Moreover, upregulation of certain gene clusters via rpoB mutation has been used in actino-…”

“…Specific gene upregulation associated with rpoB mutation has been observed in numerous model organisms, including Streptomyces spp., which are environmental organisms phylogenetically related to M. tuberculosis (35,37,69). Moreover, upregulation of certain gene clusters via rpoB mutation has been used in actino-…”