Multiple Pathway-Associated Regulations Of Nanchangmycin And Oligomycin

Streptomycetes are renowned as a rich source of natural products ofclinical, agricultural and biotechnological value. While of immensefundamental interest, insights into the regulation of antibiotic productionprovide new opportunities for knowledge-based approaches for strainimprovement to complement the classical and undoubtedly successfulstrategy of mutation and screening for improved productivity. This workfocused on the regulatory genes in nanchangmycin and oligomycinbiosynthetic clusters.There are7putative regulatory genes in nanchangmycin cluster. Two ofthem, nanR1and nanR2, encode SARP family regulators, which also sharevery high similarity with MonR1, the SARP activator of monensin. TheRT-PCR result showed that the structure genes of nan cluster composed ofeight main operons. Five of them were found consensus SARP bindingsequences within their promoter regions by genome sequence alignment. Three homolog genes of the five in mon cluster were found the sameconsensus sequence in their promoters. Disruption of nanR1or nanR2bothleaded to abolishment of nanchangmycin biosynthesis. The complementationrestored the antibiotic production. Moreover, the transcription of the structuregenes of the nan cluster were obviously decreased in the mutants. All theresults indicated that nanR1and nanR2played important roles inpathway-specific transcriptional activation of nanchangmycin cluster. Thegenetic results corroborated the bioinfomatic analysis, as the decreasedoperons were the same as the operons predicted with SARP-bindingpromoters. Meanwhile, we obtained a nanchangmycin over-producing strainby increasing the copy of nanR1-R2.Another regulatory gene nanR4encodes an AraC-family regulator.Disruption of nanR4resulted in another overproducing strain ofnanchangmycin. Complementation of this gene made the yield ofnanchangmycin recovered to the wild-type level. The transcriptional analysisof the nan cluster showed that, in the absence of NanR4, the transcription ofSARPs nanR1and nanR2were increased to4-5fold while the target genes ofSARPs were transcribed even to10-20fold. None of the other regulatorygenes in nan cluster were influenced. To conclude, NanR4was atranscriptional repressor of nanchangmycin production and it decreased nan cluster’s transcriptioin via the two SARP proteins. It is the first AraCrepressor found in the pathway of secondary metabolism.Two LAL-family regulatory genes olmRI and olmRII were found inoligomycin biosynthetic cluster. Disruption of either of them interrupted theoligomycin synthesis. When the two genes were restored, the productivity ofoligomycin in mutants were also regained. In olmRI mutant YQ6, all thestructure genes in olm cluster were decreased sharply in transcription. InolmRII mutant YQ18, only the genes located at the right flank of the clusterwere down-regulated. It gave a conclusion that OlmRI and OlmRII are bothtranscriptional activators, but their participations in the regulation aredifferent. Moreover, the abovementioned olm-deficient mutants obtained anenhanced avermectin production in Component A1a and A2a.