The LysR Transcription Factor, HexS, Is Required for Glucose Inhibition of Prodigiosin Production by Serratia marcescens

Nicholas A. Stella, James E. Fender, Roni M. Lahr, Eric J. Kalivoda, Robert M. Q. Shanks

Charles T. Campbell Laboratory of Ophthalmic Microbiology, Department of Ophthalmology, University of Pittsburgh, Pittsburgh, USA.
College of Medicine, University of Vermont, Burlington, USA.

DOI: 10.4236/aim.2012.24065                

Abstract

Generation of many useful microbe-derived secondary metabolites, including the red pigment prodigiosin of the bacterium Serratia marcescens, is inhibited by glucose. In a previous report, a genetic approach was used to determine that glucose dehydrogenase activity (GDH) is required for inhibiting prodigiosin production and transcription of the prodigiosin biosynthetic operon (pigA-N). However, the transcription factor(s) that regulate this process were not characterized. Here we tested the hypothesis that HexS, a LysR-family transcription factor similar to LrhA of Escherichia coli, is required for inhibition of prodigiosin by growth in glucose. We observed that mutation of the hexS gene in S. marcescens allowed the precocious production of prodigiosin in glucose-rich medium conditions that completely inhibited prodigiosin production by the wild type. Unlike previously described mutants able to generate prodigiosin in glucoserich medium, hexS mutants exhibited GDH activity and medium acidification similar to the wild type. Glucose inhibittion of pigA expression was shown to be dependent upon HexS, suggesting that HexS is a key transcription factor in secondary metabolite regulation in response to medium pH. These data give insight into the prodigiosin regulatory pathway and could be used to enhance the production of secondary metabolites.

Keywords

Pigment; Antibiotic; Transcription Factor; Secondary Metabolite

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1]	M. Sole, A. Francia, N. Rius and J. G. Loren, “The Role of pH in the ‘Glucose Effect’ on Prodigiosin Production by Non-Proliferating Cells of Serratia marcescens,” Letters in Applied Microbiology, Vol. 25, No. 1997, pp. 81-84.
[2]	M. Sole, N. Rius and J. G. Loren, “Rapid Extracellular Acidification Induced by Glucose Metabolism in Non-Proliferating Cells of Serratia marcescens,” International Microbiology, Vol. 3, No. 1, 2000, pp. 39-43.
[3]	N. R. Williamson, P. C. Fineran, F. J. Leeper and G. P. Salmond, “The Biosynthesis and Regulation of Bacterial Prodiginines,” Nature Reviews Microbiology, Vol. 4, No. 12, 2006, pp. 887-899. doi:10.1038/nrmicro1531
[4]	M. I. Bunting, C. F. Robinow and H. Bunting, “Factors Affecting the Elaboration of Pigment and Polysaccharide by Serratia marcescens,” Journal of Bacteriology, Vol. 58, No. 1, 1949, p. 114.
[5]	J. E. Fender, C. M. Bender, N. A. Stella, R. M. Lahr, E. J. Kalivoda and R. M. Q. Shanks, “Serratia marcescens Quinoprotein Glucose Dehydrogenase Activity Mediates Medium Acidification and Inhibition of Prodigiosin Production by Glucose,” Applied and Environmental Microbiology, Vol. 78, No. 17, 2012, pp. 6225-6235. doi:10.1128/AEM.01778-12
[6]	S. A. Dauenhauer, R. A. Hull and R. P. Williams, “Cloning and Expression in Escherichia coli of Serratia marcescens Genes Encoding Prodigiosin Biosynthesis,” Journal of Bacteriology, Vol. 158, No. 3, 1984, pp. 1128-1132.
[7]	A. K. Harris, N. R. Williamson, H. Slater, A. Cox, S. Abbasi, I. Foulds, H. T. Simonsen, F. J. Leeper and G. P. Salmond, “The Serratia Gene Cluster Encoding Biosynthesis of the Red Antibiotic, Prodigiosin, Shows Species- and Strain-Dependent Genome Context Variation,” Microbiology, Vol. 150, No. 11, 2004, pp. 3547-3560.
[8]	T. Tanikawa, Y. Nakagawa and T. Matsuyama, “Transcriptional Down Regulator HexS Controlling Prodigiosin and Serrawettin W1 Biosynthesis in Serratia marcescens,” Microbiology and Immunology, Vol. 50, No. 8, 2006, pp. 587-596.
[9]	R. M. Shanks, N. A. Stella, R. M. Lahr, S. Wang, T. I. Veverka, R. P. Kowalski and X. Liu, “Serratamolide Is a Hemolytic Factor Produced by Serratia marcescens,” PLoS One, Vol. 7, No. 5, 2012, Article ID: e36398. doi:10.1371/journal.pone.0036398
[10]	K. E. Gibson and T. J. Silhavy, “The LysR Homolog LrhA Promotes RpoS Degradation by Modulating Activity of the Response Regulator sprE,” Journal of Bacteriology, Vol. 181, No. 2, 1999, pp. 563-571.
[11]	P. C. Fineran, H. Slater, L. Everson, K. Hughes and G. P. Salmond, “Biosynthesis of Tripyrrole and Beta-Lactam Secondary Metabolites in Serratia: Integration of Quorum Sensing with Multiple New Regulatory Components in the Control of Prodigiosin and Carbapenem Antibiotic Production,” Molecular Microbiology, Vol. 56, No. 6, 2005, pp. 1495-1517. doi:10.1111/j.1365-2958.2005.04660.x
[12]	S. J. Harris, Y. L. Shih, S. D. Bentley and G. P. Salmond, “The hexA Gene of Erwinia carotovora Encodes a LysR Homologue and Regulates Motility and the Expression of Multiple Virulence Determinants,” Molecular Microbiology, Vol. 28, No. 4, 1998, pp. 705-717. doi:10.1046/j.1365-2958.1998.00825.x
[13]	A. Mukherjee, Y. Cui, W. Ma, Y. Liu and A. K. Chatterjee, “hexA of Erwinia carotovora ssp. carotovora Strain Ecc71 Negatively Regulates Production of RpoS and rsmB RNA, a Global Regulator of Extracellular Proteins, Plant Virulence and the Quorum-Sensing Signal, N-(3-oxohexanoyl)-L-homoserine Lactone,” Environmental Microbiology, Vol. 2, No. 2, 2000, pp. 203-215. doi:10.1046/j.1462-2920.2000.00093.x
[14]	S. A. Joyce and D. J. Clarke, “A hexA Homologue from Photorhabdus Regulates Pathogenicity, Symbiosis and Phenotypic Variation,” Molecular Microbiology, Vol. 47, No. 5, 2003, pp. 1445-1457. doi:10.1046/j.1365-2958.2003.03389.x
[15]	G. Bertani, “Studies on Lysogenesis. I. The Mode of Phage Liberation by Lysogenic Escherichia coli,” Journal of Bacteriology, Vol. 62, No. 3, 1951, pp. 293-300.
[16]	G. Bertani, “Lysogeny at Mid-Twentieth Century: P1, P2, and Other Experimental Systems,” Journal of Bacteriology, Vol. 186, No. 3, 2004, pp. 595-600. doi:10.1128/JB.186.3.595-600.2004
[17]	D. D. Burke D. S. Dawson and T. Stearns, “Methods in Yeast Genetics, a Cold Spring Harbor Laboratory Course Manual,” Cold Harbor Laboratory Press, Plainview, 2000.
[18]	R. M. Shanks, N. C. Caiazza, S. M. Hinsa, C. M. Toutain and G. A. O’Toole, “Saccharomyces cerevisiae-Based Molecular Tool Kit for Manipulation of Genes from Gram-Negative Bacteria,” Applied and Environmental Microbiology, Vol. 72, No. 7, 2006, pp. 5027-5036. doi:10.1128/AEM.00682-06
[19]	R. M. Shanks, D. E. Kadouri, D. P. MacEachran and A. G. O’Toole, “New Yeast Recombineering Tools for Bacteria,” Plasmid, Vol. 62, No. 2, 2009, pp. 88-97. doi:10.1016/j.plasmid.2009.05.002
[20]	E. J. Kalivoda, N. A. Stella, M. A. Aston, J. E. Fender, P. P. Thompson, R. P. Kowalski and R. M. Shanks, “Cyclic AMP Negatively Regulates Prodigiosin Production by Serratia marcescens,” Research in Microbiology, Vol. 161, No. 2, 2010, pp. 158-167. doi:10.1016/j.resmic.2009.12.004
[21]	K. Matsushita, E. Shinagawa, O. Adachi and M. Ameyama, “Quinoprotein D-Glucose Dehydrogenase of the Acinetobacter calcoaceticus Respiratory Chain: Membrane-Bound and Soluble Forms Are Different Molecular Species,” Biochemistry, Vol. 28, No. 15, 1989, pp. 6276-6280. doi:10.1021/bi00441a020
[22]	V. L. Miller and J. J. Mekalanos, “A Novel Suicide Vector and Its Use in Construction of Insertion Mutations: Osmoregulation of Outer Membrane Proteins and Virulence Determinants in Vibrio Cholerae Requires toxR,” Journal of Bacteriology, Vol. 170, 1988, p. 2575.