Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Transcription Analysis of Daptomyc in Biosynthetic Genesin Streptomyces roseosporus

  • Published : 2006.12.30
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Abstract

Insights into gene expression have the potential for improvement of antibiotic yield and the development of robust production hosts for use in recombinant biomolecule production. $Cubicin^{TM}$ (daptomycin for injection) is a recently approved antibiotic active against many Gram(+) pathogens, including those resistant to methicillin, vancomycin, and fluoroquinolones. Daptomycin is produced as a secondary metabolite by Streptomyces roseosporus. A 128 kb region of DNA including the daptomycin biosynthetic gene cluster (dpt) has been cloned. and sequenced. Using a selected array of nucleic acid probes representing this region, we compared the expression levels of the dpt genes between S. roseosporus wild-type (WT) and derived S. roseosporus high-producer of daptomycin (HP). We observed that the majority of the biosynthetic genes were upregulated in HP compared with WT; a total of 12 genes, including those encoding daptomycin synthetase, showed consistently and significantly higher expression levels, at least 5-fold, in HP compared with WT. In contrast, some genes, flanking the dpt cluster, were expressed at higher levels in the WT strain. The expression of housekeeping genes such as S. roseosporus rpsL, rpsG, and 16S (positive controls) and presumptive intergenic regions in the dpt cluster (negative control) were identical in the two strains. In addition, we compared transcription during the early, mid-log, and early-stationary phases of growth in the HP strain. The same set of genes was upregulated and downregulated under all conditions examined; housekeeping genes showed no relative change in expression level over the periods of growth tested. Analyses of this type would be of value in studies of strain improvement and also for the identification of gene regulation processes that are important for secondary metabolite production.

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References

  1. Baltz, R. H. 1997. Lipopeptide antibiotics produced by Streptomyces roseosporus and Streptomyces fradiae, pp. 415-435. In W. R. Strohl (ed.). Biotechnology of Antibiotics, 2nd Ed. Marcel Dekker, Inc., New York
  2. Borovok, I., R. Kreisberg-Zakarin, M. Yanko, R. Schreiber, M. Myslovati, F. Aslund, A. Holmgren, G. Cohen, and Y. Aharonowitz. 2002. Streptomyces spp. contain class Ia and class II ribonucleotide reductases: Expression analysis of the vegetative growth. Microbiology 148: 391-404 https://doi.org/10.1099/00221287-148-2-391
  3. Bucca, G. B., A. M. Brassington, G. Hotchkiss, V. Mersinias, and C. P. Smith. 2003. Negative feedback regulation of DnaK, clpB, and Ion expression by the DnaK chaperone machine in Streptomyces coelicolor, identified by transcriptome and in vivo DnaK-depletion analysis. Mol. Microbiol. 50: 153-166 https://doi.org/10.1046/j.1365-2958.2003.03696.x
  4. Bush, L., J. A. M. Boscia, and D. Kaye. 1998. Daptomycin (LY146032) treatment of experimental Enterococcal endocarditis. Antimicrob. Agents Chemother. 32: 877-881
  5. David, Y., C. W. James, and G. R. Jose-Carlos. 2001. Microarray for studying the host transcriptional response to microbial infection and for the identification of host drug targets. Microb. Infect. 3: 813-821 https://doi.org/10.1016/S1286-4579(01)01439-3
  6. Debono, M., B. J. Abbott, R. M. Molly, D. S. Fukuda, A. H. Hunt, V. M. Daupert, F. T. Counter, J. L. Ott, C. B. Carrell, and L. C. Howard. 1988. Enzymatic and chemical modifications of lipopeptide antibiotic A21978C: The synthesis and evaluation of daptomycin (LY146032). J. Antibiot. (Tokyo) 41: 1093-1105 https://doi.org/10.7164/antibiotics.41.1093
  7. Debono, M., C. B. Barnhart, J. A. Carrell, J. L. Hoffman, B. J. Occolowitz, B. J. Abbott, D. S. Fukuda, and R. L. Hamill. 1987. A21978C, a complex of new acidic peptide antibiotics: Isolation, chemistry, and mass spectral structure elucidation. J. Antibiot. 40: 761-777 https://doi.org/10.7164/antibiotics.40.761
  8. Demain, A. L. and J. E. Davies. 1999. Manual of Industrial Microbiology and Biotechnology. ASM Press, Washington, D.C
  9. Garrison, M. W., K. Vance-Bryan, T. A. Larson, J. P. Toscano, and J. C. Rotschafer. 1990. Assessment of effects of protein binding on daptomycin and vancomycin killing of Staphylococcus aureus by using an in vitro pharmacodynamic model. Antimicrob. Agents Chemother. 31: 625-629
  10. Huang, J., C. J. Lih, K. H. Pan, and S. N. Cohen. 2001. Global analysis of growth phase responsive gene expression and regulation of antibiotic biosynthetic pathways in Streptomyces coelicolor using DNA microarray. Gene Develop. 15: 3183-3192 https://doi.org/10.1101/gad.943401
  11. Hu, H. and K. Ochi. 2001. Novel approach for improving the productivity of antibiotic-producing strains by inducing combined resistant mutations. Appl. Environ. Microbiol. 67: 1885-1892 https://doi.org/10.1128/AEM.67.4.1885-1892.2001
  12. Kim, S., J. S. Kang, D. D. Jang, K. K. Lee, S. A. Kim, B. S. Han, and Y. I. Park. 2004. Differential gene expression in estradiol-3-benzoate-treated liver and chemically-induced hepatocellular carcinoma. J. Microbiol. Biotechnol. 14: 1286- 1294
  13. Lal, R., R. Khanna, H. Kaur, M. Khanna, N. Dhingra, S. Lal, K. H. Gartemann, R. Eichenlaub, and P. K. Ghosh. 1996. Engineering antibiotic producers to overcome the limitations of classical strain improvement programs. Crit. Rev. Microbiol. 22: 201-255 https://doi.org/10.3109/10408419609105481
  14. Lee, J. Y. and N. G. Lee. 2004. Transcriptional responses of human respiratory epithelial cells to nontypeable Haemophilus influenzae infection analyzed by high density cDNA microarrays. J. Microbiol. Biotechnol. 14: 836-843
  15. Lee, M. L., F. C. Kuo, G. A. Whitmore, and J. Sklar. 2000. Importance of replication in microarray gene expression studies: Statistical methods and evidence from repetitive cDNA hybridizations. Proc. Nat. Acad. Sci. USA 97: 9834- 9839
  16. Lee, S. H. and Y. T. Rho. 1999. Improvement of tylosin fermentation by mutation and medium optimization. Lett. Appl. Microbiol. 28: 142-144 https://doi.org/10.1046/j.1365-2672.1999.00478.x
  17. Miao, V., M.-F. Coeffet-Le Gal, P. Brian, R. Brost, J. Penn, A. Whiting, S. Martin, R. Ford, R. Parr, M. Bouchard, C. J. Silva, S. K. Wrigley, and R. H. Baltz. 2005. Daptomycin biosynthesis in Streptomyces roseosporus: Cloning and analysis of the gene cluster and revision of peptide stereochemistry. Microbiology 151: 1507-1523 https://doi.org/10.1099/mic.0.27757-0
  18. McHenney, M. A., T. J. Hosted, B. S. Dehoff, P. R. Rosteck, and R. H. Baltz. 1998. Molecular cloning and physical mapping of the daptomycin gene cluster from Streptomyces roseosporus. J. Bacteriol. 180: 143-151
  19. McHenney, M. A. and R. H. Baltz. 1996. Gene transfer and transposition mutagenesis in Streptomyces roseosporus: Mapping of insertions that influence daptomycin or pigment production. Microbiology 142: 2363-2373 https://doi.org/10.1099/00221287-142-9-2363
  20. Nelson, C. C., D. Hoffart, E. M. Gleave, and P. S. Rennie. 2003. Application of gene microarray in the study of prostate cancer. Methods Mol. Med. 81: 299-320
  21. Oh, K. S., O. S. Kwon, Y. W. Oh, M. J. Sohn, S. G. Jung, Y. K. Kim, M. G. Kim, S. K. Rhee, G. Gellissen, and H. A. Kang. 2004. Fabrication of partial genome microarray of the methylotrophic yeast Hansenula polymorpha: Optimization and evaluation of transcript profiling. J. Microbiol. Biotechnol. 14: 1239-1248
  22. Ryding N. J., T. B. Anderson, and W. C. Champness. 2002. Regulation of the Streptomyces coelicolor calciumdependent antibiotic by absA, encoding a cluster-linked twocomponent system. J. Bacteriol. 184: 794-805 https://doi.org/10.1128/JB.184.3.794-805.2002
  23. Rybak, M. J., E. Hershberger, E. T. Moldovan, and R. G. Grucz. 2000. In vitro activities of daptomycin, vancomycin, linezolid, and quinopristin-dalfopristin against Staphylococci and Enterococci, including vancomycin intermediate and resistant strains. Antimicrob. Agents Chemother. 34: 2081- 2085
  24. Sambrook, J., E. F. Fritsch, and T. Maniatis. 1989. Molecular Cloning: A Laboratory Manual, 2nd Ed. Cold Spring Harbor Laboratory Press. Cold Spring Harbor, N.Y
  25. Seto, M., E. Masai, M. Ida, T. Hatta, K. Kimbara, M. Fukuda, and K. Yano. 1995. Multiple polychlorinated biphenyl transformation systems in the gram-positive bacterium Rhodococcus sp. strain RHA1. Appl. Environ. Microbiol. 61: 4510-4513
  26. Takano, E., H. C. Gramajo, E. Strauch, N. Andres, J. White, and M. J. Bibb. 1992. Transcriptional regulation of the redD transcriptional activator gene accounts for growth-phase dependent production of the antibiotic undecylprodigiosin in Streptomyces coelicolor A3(2). Mol. Microbiol. 6: 2797- 2804 https://doi.org/10.1111/j.1365-2958.1992.tb01459.x
  27. Wagner, V. E., D. Bushnell, L. Passador, A. I. Brooks, and B. H. Iglewski. 2003. Microarray analysis of Pseudomonas aeruginosa quorum-sensing regulons: Effects of growth phase and environment. J. Bacteriol. 185: 2080-2095 https://doi.org/10.1128/JB.185.7.2080-2095.2003
  28. Wang, H. Y., L. M. Renae, E. K. Anne, S. G. Andrew, V. L. Truong, B. Babak, F. Bryan, Q. John, and N. H. Lee. 2003. Assessing unmodified 70-mer oligonucleotide probe performance on glass-slide microarrays. Genome Biol. 4: R5 https://doi.org/10.1186/gb-2003-4-10-r70
  29. Zhang, Y. X., K. Perry, V. A. Vinci, K. Powell, W. P. Stemmer, and S. B. del Cardayre. 2002. Genome shuffling leads to rapid phenotype improvement in bacteria. Nature 415: 644-646 https://doi.org/10.1038/415644a
  30. Zhao, X., S. Nampalli, A. J. Serino, and S. Kumar. 2001. Immobilization of oligonucleotides with multiple anchors to microchips. Nucleic Acids Res. 29: 955-959 https://doi.org/10.1093/nar/29.4.955