1887

Abstract

ARC2 is a synthetic compound, related in structure and mechanism to the antibiotic triclosan, that activates the production of many specialized metabolites in the genus of bacteria. In this work, we demonstrate that the addition of ARC2 to cultures results in considerable alterations in overall gene expression including most notably the specialized metabolic genes. Using actinorhodin production as a model system, we show that the effect of ARC2 depends on the pleiotropic regulators and but not . We find that the constitutive expression of can bypass the need for but not the reverse, while the constitutive expression of had no effect on actinorhodin production. These data are consistent with a model in which ARC2 activates a cell stress response that depends on AfsR activating the expression of the gene such that AfsS then triggers the production of actinorhodin.

Funding
This study was supported by the:
  • Canadian Institutes of Health Research (Award MOP-133636)
    • Principle Award Recipient: JustinRea Nodwell
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/content/journal/micro/10.1099/mic.0.001047
2021-05-04
2021-05-15
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References

  1. Raja A, Prabakaran P. Actinomycetes and drug-an overview. Am J Drug Discov Dev 2011; 1:75–84 [CrossRef]
    [Google Scholar]
  2. van Keulen G, Dyson PJ. Production of specialized metabolites by Streptomyces coelicolor A3(2). Adv Appl Microbiol 2014; 89:217-66 [CrossRef][PubMed]
    [Google Scholar]
  3. Breukink E, de Kruijff B. Lipid II as a target for antibiotics. Nat Rev Drug Discov 2006; 5:321–323 [CrossRef][PubMed]
    [Google Scholar]
  4. Campbell EA, Korzheva N, Mustaev A, Murakami K, Nair S. Structural mechanism for rifampicin inhibition of bacterial RNA polymerase. Cell 2001; 104:901–912 [CrossRef][PubMed]
    [Google Scholar]
  5. Chen CR, Malik M, Snyder M, Drlica K. Dna gyrase and topoisomerase IV on the bacterial chromosome: quinolone-induced DNA cleavage. J Mol Biol 1996; 258:627–637 [CrossRef][PubMed]
    [Google Scholar]
  6. Kohanski MA, Dwyer DJ, Collins JJ. How antibiotics kill bacteria: from targets to networks. Nat Rev Microbiol 2010; 8:423–435 [CrossRef][PubMed]
    [Google Scholar]
  7. Vázquez-Laslop N, Mankin AS. How macrolide antibiotics work. Trends Biochem Sci 2018; 43:668–684 [CrossRef][PubMed]
    [Google Scholar]
  8. Ho LK, Nodwell JR. David and Goliath: chemical perturbation of eukaryotes by bacteria. J Ind Microbiol Biotechnol 2016; 43:233–248 [CrossRef][PubMed]
    [Google Scholar]
  9. Aslam B, Wang W, Arshad MI, Khurshid M, Muzammil S. Antibiotic resistance: a rundown of a global crisis. Infect Drug Resist 2018; 11:1645–1658 [CrossRef][PubMed]
    [Google Scholar]
  10. Brown ED, Wright GD. Antibacterial drug discovery in the resistance era. Nature 2016; 529:336–343 [CrossRef][PubMed]
    [Google Scholar]
  11. Craney A, Ahmed S, Nodwell J. Towards a new science of secondary metabolism. J Antibiot 2013; 66:387–400 [CrossRef][PubMed]
    [Google Scholar]
  12. Daniel-Ivad M, Pimentel-Elardo S, Nodwell JR. Control of specialized metabolism by signaling and transcriptional regulation: opportunities for new platforms for drug discovery?. Annu Rev Microbiol 2018; 72:25–48 [CrossRef][PubMed]
    [Google Scholar]
  13. Yoon V, Nodwell JR. Activating secondary metabolism with stress and chemicals. J Ind Microbiol Biotechnol 2014; 41:415–424 [CrossRef]
    [Google Scholar]
  14. Zhang X, Elliot MA. Unlocking the trove of metabolic treasures: activating silent biosynthetic gene clusters in bacteria and fungi. Curr Opin Microbiol 2019; 51:9–15 [CrossRef][PubMed]
    [Google Scholar]
  15. Mak S, Nodwell JR. Actinorhodin is a redox-active antibiotic with a complex mode of action against gram-positive cells. Mol Microbiol 2017; 106:597–613 [CrossRef][PubMed]
    [Google Scholar]
  16. Brown ED. Screening in academe: a perspective on implementation of university-based small molecule screening. J Biomol Screen 2003; 8:377–379 [CrossRef][PubMed]
    [Google Scholar]
  17. Craney A, Ozimok C, Pimentel-Elardo SM, Capretta A, Nodwell JR. Chemical perturbation of secondary metabolism demonstrates important links to primary metabolism. Chem Biol 2012; 19:1020–1027 [CrossRef][PubMed]
    [Google Scholar]
  18. Heath RJ, Rubin JR, Holland DR, Zhang E, Snow ME et al. Mechanism of triclosan inhibition of bacterial fatty acid synthesis. J Biol Chem 1999; 274:11110–11114 [CrossRef][PubMed]
    [Google Scholar]
  19. Heath RJ, Rock CO. Enoyl-acyl carrier protein reductase (fabI) plays a determinant role in completing cycles of fatty acid elongation in Escherichia coli . J Biol Chem 1995; 270:26538–26542 [CrossRef]
    [Google Scholar]
  20. Ahmed S, Craney A, Pimentel-Elardo SM, Nodwell JR. A synthetic, species-specific activator of secondary metabolism and sporulation in Streptomyces coelicolor . Chembiochem 2013; 14:83–91 [CrossRef][PubMed]
    [Google Scholar]
  21. Pimentel-Elardo SM, Sørensen D, Ho L, Ziko M, Bueler SA. Activity-Independent discovery of secondary metabolites using chemical elicitation and cheminformatic inference. ACS Chem Biol 2015; 10:2616–2623 [CrossRef][PubMed]
    [Google Scholar]
  22. Lee PC, Umeyama T, Horinouchi S. afsS is a target of AfsR, a transcriptional factor with ATPase activity that globally controls secondary metabolism in Streptomyces coelicolor A3(2). Mol Microbiol 2002; 43:1413–1430 [CrossRef][PubMed]
    [Google Scholar]
  23. Matsumoto A, Hong SK, Ishizuka H, Horinouchi S, Beppu T. Phosphorylation of the AfsR protein involved in secondary metabolism in Streptomyces species by a eukaryotic-type protein kinase. Gene 1994; 146:47–56 [CrossRef][PubMed]
    [Google Scholar]
  24. Cobb RE, Wang Y, Zhao H. High-Efficiency multiplex genome editing of Streptomyces species using an engineered CRISPR/Cas system. ACS Synth Biol 2015; 4:723–728 [CrossRef][PubMed]
    [Google Scholar]
  25. Tong Y, Charusanti P, Zhang L, Weber T, Lee SY. Crispr-Cas9 based engineering of Actinomycetal genomes. ACS Synth Biol 2015; 4:1020–1029 [CrossRef][PubMed]
    [Google Scholar]
  26. Kieser T, Bibb MJ, Buttner MJ, Chater KF. Practical Streptomyces Genetics.. In John Innes Foundation, 2nd ed. 2000
    [Google Scholar]
  27. Kirby KS, Fox-Carter E, Guest M. Isolation of deoxyribonucleic acid and ribosomal ribonucleic acid from bacteria. Biochem J 1967; 104:258–262 [CrossRef][PubMed]
    [Google Scholar]
  28. Bolger AM, Lohse M, Usadel B. Trimmomatic: a flexible trimmer for illumina sequence data. Bioinformatics 2014; 30:2114–2120 [CrossRef][PubMed]
    [Google Scholar]
  29. Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nat Methods 2012; 9:357–359 [CrossRef]
    [Google Scholar]
  30. Anders S, Pyl PT, Huber W. HTSeq--a Python framework to work with high-throughput sequencing data. Bioinformatics 2015; 31:166–169 [CrossRef][PubMed]
    [Google Scholar]
  31. Love MI, Huber W, Anders S. Moderated estimation of fold change and dispersion for RNA-Seq data with DESeq2. Genome Biol 2014; 15:550 [CrossRef][PubMed]
    [Google Scholar]
  32. Huerta-Cepas J, Szklarczyk D, Forslund K, Cook H, Heller D. eggNOG 4.5: a hierarchical orthology framework with improved functional annotations for eukaryotic, prokaryotic and viral sequences. Nucleic Acids Res 2016; 44:D286–D293 [CrossRef]
    [Google Scholar]
  33. Craney A, Hohenauer T, Xu Y, Navani NK, Li Y et al. A synthetic luxCDABE gene cluster optimized for expression in high-GC bacteria. Nucleic Acids Res 2007; 35:e46 [CrossRef][PubMed]
    [Google Scholar]
  34. Bierman M, Logan R, O'Brien K, Seno ET, Rao RN et al. Plasmid cloning vectors for the conjugal transfer of DNA from Escherichia coli to Streptomyces spp. Gene 1992; 116:43–49 [CrossRef][PubMed]
    [Google Scholar]
  35. Bursy J, Kuhlmann AU, Pittelkow M, Hartmann H, Jebbar M et al. Synthesis and uptake of the compatible solutes ectoine and 5-hydroxyectoine by Streptomyces coelicolor A3(2) in response to salt and heat stresses. Appl Environ Microbiol 2008; 74:7286–7296 [CrossRef][PubMed]
    [Google Scholar]
  36. Hojati Z, Milne C, Harvey B, Gordon L, Borg M et al. Structure, biosynthetic origin, and engineered biosynthesis of calcium-dependent antibiotics from Streptomyces coelicolor . Chem Biol 2002; 9:1175–1187 [CrossRef][PubMed]
    [Google Scholar]
  37. Wright LF, Hopwood DA. Actinorhodin is a chromosomally-determined antibiotic in Streptomyces coelicolar A3(2). J Gen Microbiol 1976; 96:289–297 [CrossRef][PubMed]
    [Google Scholar]
  38. Feitelson JS, Malpartida F, Hopwood DA. Genetic and biochemical characterization of the red gene cluster of Streptomyces coelicolor A3(2). J Gen Microbiol 1985; 131:2431–2441 [CrossRef][PubMed]
    [Google Scholar]
  39. Takano E, Nihira T, Hara Y, Jones JJ, Gershater CJL et al. Purification and structural determination of SCB1, a γ-butyrolactone That elicits antibiotic production Streptomyces coelicolor A3(2). J Biol Chem 2000; 275:11010–11016 [CrossRef]
    [Google Scholar]
  40. Gomez-Escribano JP, Song L, Fox DJ, Yeo V, Bibb MJ et al. Structure and biosynthesis of the unusual polyketide alkaloid coelimycin P1, a metabolic product of the cpk gene cluster of Streptomyces coelicolor M145. Chemical Science 2012; 3:2716–2720 [CrossRef]
    [Google Scholar]
  41. Poralla K, Muth G, Härtner T. Hopanoids are formed during transition from substrate to aerial hyphae in Streptomyces coelicolor A3(2). FEMS Microbiol Lett 2000; 189:93–95 [CrossRef][PubMed]
    [Google Scholar]
  42. Cruz-Morales P, Kopp JF, Martínez-Guerrero C, Yáñez-Guerra LA, Selem-Mojica N et al. Phylogenomic analysis of natural products biosynthetic gene clusters allows discovery of arseno-organic metabolites in model streptomycetes. Genome Biol Evol 2016; 8:1906–1916 [CrossRef][PubMed]
    [Google Scholar]
  43. Nett M, Ikeda H, Moore BS. Genomic basis for natural product biosynthetic diversity in the actinomycetes. Nat Prod Rep 2009; 26:1362–1384 [CrossRef][PubMed]
    [Google Scholar]
  44. Song L, Barona-Gómez F, Corre C, Xiang L, Udwary DW et al. Type III polyketide synthase beta-ketoacyl-ACP starter unit and ethylmalonyl-CoA extender unit selectivity discovered by Streptomyces coelicolor genome mining. J Am Chem Soc 2006; 128:14754–14755 [CrossRef][PubMed]
    [Google Scholar]
  45. Bentley SD, Chater KF, Cerdeño-Tárraga AM, Challis GL, Thomson NR. Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2). Nature 2002; 417:141–147 [CrossRef]
    [Google Scholar]
  46. Takano H, Obitsu S, Beppu T, Ueda K. Light-induced carotenogenesis in Streptomyces coelicolor A3(2): identification of an extracytoplasmic function sigma factor that directs photodependent transcription of the carotenoid biosynthesis gene cluster. J Bacteriol 2005; 187:1825–1832 [CrossRef][PubMed]
    [Google Scholar]
  47. Kodani S, Hudson ME, Durrant MC, Buttner MJ, Nodwell JR et al. The SapB morphogen is a lantibiotic-like peptide derived from the product of the developmental gene ramS in Streptomyces coelicolor . Proc Natl Acad Sci U S A 2004; 101:11448–11453 [CrossRef][PubMed]
    [Google Scholar]
  48. Ma P, Patching SG, Ivanova E, Baldwin JM, Sharples D et al. Allantoin transport protein, Pucl, from Bacillus subtilis: Evolutionary relationships, amplified expression, activity and specificity. Microbiology 2016
    [Google Scholar]
  49. Navone L, Casati P, Licona-Cassani C, Marcellin E, Nielsen LK. Allantoin catabolism influences the production of antibiotics in Streptomyces coelicolor . Appl Microbiol Biotechnol 2014; 98:351–360 [CrossRef][PubMed]
    [Google Scholar]
  50. Navone L, Macagno JP, Licona-Cassani C, Marcellin E, Nielsen LK. AllR controls the expression of Streptomyces coelicolor allantoin pathway genes. Appl Environ Microbiol 2015; 81:6649–6659 [CrossRef][PubMed]
    [Google Scholar]
  51. Bibb MJ, Molle V, Buttner MJ. sigma(BldN), an extracytoplasmic function RNA polymerase sigma factor required for aerial mycelium formation in Streptomyces coelicolor A3(2). J Bacteriol 2000; 182:4606–4616 [CrossRef][PubMed]
    [Google Scholar]
  52. Champness WC. New loci required for Streptomyces coelicolor morphological and physiological differentiation. J Bacteriol 1988; 170:1168–1174 [CrossRef][PubMed]
    [Google Scholar]
  53. Gehring AM, Yoo NJ, Losick R. RNA polymerase sigma factor that blocks morphological differentiation by Streptomyces coelicolor . J Bacteriol 2001; 183:5991–5996 [CrossRef][PubMed]
    [Google Scholar]
  54. Chater KF. Regulation of sporulation in Streptomyces coelicolor A3(2): a checkpoint multiplex?. Curr Opin Microbiol 2001; 4:667–673 [CrossRef][PubMed]
    [Google Scholar]
  55. Mendez C, Chater KF. Cloning of whiG, a gene critical for sporulation of Streptomyces coelicolor A3(2). J Bacteriol 1987; 169:5715–5720 [CrossRef][PubMed]
    [Google Scholar]
  56. O'Connor TJ, Kanellis P, Nodwell JR. The ramC gene is required for morphogenesis in Streptomyces coelicolor and expressed in a cell type-specific manner under the direct control of RamR. Mol Microbiol 2002; 45:45–57 [CrossRef][PubMed]
    [Google Scholar]
  57. Floriano B, Bibb M. afsR is a pleiotropic but conditionally required regulatory gene for antibiotic production in Streptomyces coelicolor A3(2). Mol Microbiol 1996; 21:385–396 [CrossRef][PubMed]
    [Google Scholar]
  58. Horinouchi S. AfsR as an integrator of signals that are sensed by multiple serine/threonine kinases in Streptomyces coelicolor A3(2). J Ind Microbiol Biotechnol 2003; 30:462–467 [CrossRef][PubMed]
    [Google Scholar]
  59. Matsumoto A, Ishizuka H, Beppu T, Horinouchi S. Involvement of a small ORF downstream of the afsR gene in the regulation of secondary metabolism in Streptomyces coelicolor A3(2). Actinomycetologica 1995; 9:37–43 [CrossRef]
    [Google Scholar]
  60. Tomono A, Mashiko M, Shimazu T, Inoue H, Nagasawa H et al. Self-Activation of serine/threonine kinase AfsK on autophosphorylation at threonine-168. J Antibiot 2006; 59:117–123 [CrossRef][PubMed]
    [Google Scholar]
  61. Tanaka A, Takano Y, Ohnishi Y, Horinouchi S. AfsR recruits RNA polymerase to the afsS promoter: a model for transcriptional activation by SARPs. J Mol Biol 2007; 369:322–333 [CrossRef]
    [Google Scholar]
  62. Hempel AM, Cantlay S, Molle V, Wang SB, Naldrett MJ et al. The ser/thr protein kinase AfsK regulates polar growth and hyphal branching in the filamentous bacteria Streptomyces.. Proceedings of the National Academy of Sciences of the United States of America 2012
    [Google Scholar]
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