Repression of Antibiotic Production and Sporulation in Streptomyces coelicolor by Overexpression of a TetR Family Transcriptional Regulator

ABSTRACT The overexpression of a regulatory gene of the TetR family (SCO3201) originating either from Streptomyces lividans or from Streptomyces coelicolor was shown to strongly repress antibiotic production (calcium-dependent antibiotic [CDA], undecylprodigiosin [RED], and actinorhodin [ACT]) of S. coelicolor and of the ppk mutant strain of S. lividans. Curiously, the overexpression of this gene also had a strong inhibitory effect on the sporulation process of S. coelicolor but not on that of S. lividans. SCO3201 was shown to negatively regulate its own transcription, and its DNA binding motif was found to overlap its −35 promoter sequence. The interruption of this gene in S. lividans or S. coelicolor did not lead to any obvious phenotypes, indicating that when overexpressed SCO3201 likely controls the expression of target genes of other TetR regulators involved in the regulation of the metabolic and morphological differentiation process in S. coelicolor. The direct and functional interaction of SCO3201 with the promoter region of scbA, a gene under the positive control of the TetR-like regulator, ScbR, was indeed demonstrated by in vitro as well as in vivo approaches.

[1]  C. Choi,et al.  Modulation of Actinorhodin Biosynthesis in Streptomyces lividans by Glucose Repression of afsR2 Gene Transcription , 2001, Journal of bacteriology.

[2]  Charles Elkan,et al.  Fitting a Mixture Model By Expectation Maximization To Discover Motifs In Biopolymer , 1994, ISMB.

[3]  J. Walker,et al.  Over-production of proteins in Escherichia coli: mutant hosts that allow synthesis of some membrane proteins and globular proteins at high levels. , 1996, Journal of molecular biology.

[4]  R. Kirby,et al.  Genetic determination of methylenomycin synthesis by the SCP1 plasmid of Streptomyces coelicolor A3(2). , 1977, Journal of general microbiology.

[5]  C. Thompson,et al.  Cloning and expression of the tyrosinase gene from Streptomyces antibioticus in Streptomyces lividans. , 1983, Journal of general microbiology.

[6]  S. Horinouchi,et al.  A microbial hormone, A-factor, as a master switch for morphological differentiation and secondary metabolism in Streptomyces griseus. , 2002, Frontiers in bioscience : a journal and virtual library.

[7]  Sarika Mehra,et al.  A Bistable Gene Switch for Antibiotic Biosynthesis: The Butyrolactone Regulon in Streptomyces coelicolor , 2008, PloS one.

[8]  M. Schell,et al.  Footprinting with an automated capillary DNA sequencer. , 2000, BioTechniques.

[9]  A. Pühler,et al.  A vector system with temperature-sensitive replication for gene disruption and mutational cloning in streptomycetes , 1989, Molecular and General Genetics MGG.

[10]  P. Youngman,et al.  xylE functions as an efficient reporter gene in Streptomyces spp.: use for the study of galP1, a catabolite-controlled promoter , 1989, Journal of bacteriology.

[11]  K. Chater,et al.  Genetics of differentiation in Streptomyces. , 1993, Annual review of microbiology.

[12]  Gregory L. Challis,et al.  Synergy and contingency as driving forces for the evolution of multiple secondary metabolite production by Streptomyces species , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[13]  M. Bibb,et al.  Deletion of a regulatory gene within the cpk gene cluster reveals novel antibacterial activity in Streptomyces coelicolor A3(2). , 2010, Microbiology.

[14]  P. Bruheim,et al.  High-yield actinorhodin production in fed-batch culture by a Streptomyces lividans strain overexpressing the pathway-specific activator gene actII-ORF4 , 2002, Journal of Industrial Microbiology and Biotechnology.

[15]  D. Hopwood,et al.  Genetic and biochemical characterization of the red gene cluster of Streptomyces coelicolor A3(2). , 1985, Journal of general microbiology.

[16]  M. Buttner,et al.  Sensing and responding to diverse extracellular signals? Analysis of the sensor kinases and response regulators of Streptomyces coelicolor A3(2). , 2004, Microbiology.

[17]  J. Suh,et al.  Accumulation of S-Adenosyl-l-Methionine Enhances Production of Actinorhodin but Inhibits Sporulation in Streptomyces lividans TK23 , 2003, Journal of bacteriology.

[18]  Y. Yamada,et al.  A complex role for the gamma-butyrolactone SCB1 in regulating antibiotic production in Streptomyces coelicolor A3(2). , 2001, Molecular microbiology.

[19]  K. Chater,et al.  A developmentally regulated gene encoding a repressor‐like protein is essential for sporulation in Streptomyces coelicolor A3(2) , 1998, Molecular microbiology.

[20]  K. Ochi,et al.  Activation of Antibiotic Biosynthesis by Specified Mutations in the rpoB Gene (Encoding the RNA Polymerase β Subunit) of Streptomyces lividans , 2002, Journal of bacteriology.

[21]  Jason Micklefield,et al.  Structure, biosynthetic origin, and engineered biosynthesis of calcium-dependent antibiotics from Streptomyces coelicolor. , 2002, Chemistry & biology.

[22]  H. Chouayekh,et al.  The polyphosphate kinase plays a negative role in the control of antibiotic production in Streptomyces lividans , 2002, Molecular microbiology.

[23]  T. Kieser.  Practical streptomyces genetics , 2000 .

[24]  H. Ogawara,et al.  Expression and Characterization of the Streptomyces coelicolor Serine/Threonine Protein Kinase PkaD , 2008, Bioscience, biotechnology, and biochemistry.

[25]  L. Serrano,et al.  Engineering stability in gene networks by autoregulation , 2000, Nature.

[26]  D. Glover.  DNA cloning : a practical approach , 1985 .

[27]  K. Ochi,et al.  Increased expression of ribosome recycling factor is responsible for the enhanced protein synthesis during the late growth phase in an antibiotic‐overproducing Streptomyces coelicolor ribosomal rpsL mutant , 2006, Molecular microbiology.

[28]  S. Horinouchi,et al.  Involvement of a Small ORF Downstream of the afsR Gene in the Regulation of Secondary Metabolism in Streptomyces coelicolor A3(2). , 1995 .

[29]  C. W. Chen,et al.  The cutRS signal transduction system of Streptomyces lividans represses the biosynthesis of the polyketide antibiotic actinorhodin. , 1996, Molecular microbiology.

[30]  J. Kormanec,et al.  Transcriptional Studies and Regulatory Interactions between the phoR-phoP Operon and the phoU, mtpA, and ppk Genes of Streptomyces lividans TK24 , 2006, Journal of bacteriology.

[31]  Stanley N Cohen,et al.  afsR2: a previously undetected gene encoding a 63‐amino‐acid protein that stimulates antibiotic production in Streptomyces lividans , 1994, Molecular microbiology.

[32]  R. Rappuoli,et al.  The Iron-Responsive Regulator Fur Is Transcriptionally Autoregulated and Not Essential in Neisseria meningitidis , 2003, Journal of bacteriology.

[33]  D. Aceti,et al.  Transcriptional Regulation of Streptomyces coelicolorPathway-Specific Antibiotic Regulators by the absA andabsB Loci , 1998, Journal of bacteriology.

[34]  Klas Flärdh,et al.  Streptomyces morphogenetics: dissecting differentiation in a filamentous bacterium , 2009, Nature Reviews Microbiology.

[35]  S. Oliver,et al.  Dispersed growth of Streptomyces in liquid culture , 1989, Applied Microbiology and Biotechnology.

[36]  Claudia Sala,et al.  Mycobacterium tuberculosis FurA Autoregulates Its Own Expression , 2003, Journal of bacteriology.

[37]  Raquel Tobes,et al.  The TetR Family of Transcriptional Repressors , 2005, Microbiology and Molecular Biology Reviews.

[38]  S. Horinouchi,et al.  Involvement of two A‐factor receptor homologues in Streptomyces coelicolor A3(2) in the regulation of secondary metabolism and morphogenesis , 1998, Molecular microbiology.

[39]  S. Horinouchi,et al.  Autorepression of AdpA of the AraC/XylS family, a key transcriptional activator in the A-factor regulatory cascade in Streptomyces griseus. , 2005, Journal of molecular biology.

[40]  M. Bibb,et al.  A response-regulator-like activator of antibiotic synthesis from Streptomyces coelicolor A3(2) with an amino-terminal domain that lacks a phosphorylation pocket. , 1998, Microbiology.

[41]  J. Martín,et al.  The two-component PhoR-PhoP system controls both primary metabolism and secondary metabolite biosynthesis in Streptomyces lividans , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[42]  S. Baumberg,et al.  Transcriptional activation of the pathway‐specific regulator of the actinorhodin biosynthetic genes in Streptomyces coelicolor , 2005, Molecular microbiology.

[43]  F. Studier,et al.  Use of T7 RNA polymerase to direct expression of cloned genes. , 1990, Methods in enzymology.

[44]  S. Horinouchi,et al.  An AfsK/AfsR system involved in the response of aerial mycelium formation to glucose in Streptomyces griseus. , 1999, Microbiology.

[45]  M. Bibb,et al.  The regulation of antibiotic production in Streptomyces coelicolor A3(2) , 1996 .

[46]  Eriko Takano,et al.  A bacterial hormone (the SCB1) directly controls the expression of a pathway‐specific regulatory gene in the cryptic type I polyketide biosynthetic gene cluster of Streptomyces coelicolor , 2005, Molecular microbiology.

[47]  Guoping Zhao,et al.  Characterization of rrdA, a TetR Family Protein Gene Involved in the Regulation of Secondary Metabolism in Streptomyces coelicolor , 2009, Applied and Environmental Microbiology.

[48]  C. Thompson,et al.  Gene cloning in Streptomyces. , 1982, Current topics in microbiology and immunology.

[49]  J. White,et al.  bldA dependence of undecylprodigiosin production in Streptomyces coelicolor A3(2) involves a pathway-specific regulatory cascade , 1997, Journal of bacteriology.

[50]  Stanley N Cohen,et al.  Putative TetR Family Transcriptional Regulator SCO1712 Encodes an Antibiotic Downregulator in Streptomyces coelicolor , 2010, Applied and Environmental Microbiology.

[51]  P. Branny,et al.  Antibiotic resistance gene cassettes derived from the omega interposon for use in E. coli and Streptomyces. , 1997, Gene.

[52]  M. Bibb,et al.  Cloning and analysis of the promoter region of the erythromycin resistance gene (ermE) of Streptomyces erythraeus. , 1985, Gene.

[53]  Eriko Takano,et al.  A cryptic type I polyketide synthase (cpk) gene cluster in Streptomyces coelicolor A3(2) , 2007, Archives of Microbiology.

[54]  E. Takano,et al.  Transcriptional regulation of the redD transcriptional activator gene accounts for growth‐phase‐dependent production of the antibiotic undecylprodigiosin in Streptomyces coelicolor A3(2) , 1992, Molecular microbiology.

[55]  J. Nodwell,et al.  Phosphorylated AbsA2 Negatively Regulates Antibiotic Production in Streptomyces coelicolor through Interactions with Pathway-Specific Regulatory Gene Promoters , 2007, Journal of bacteriology.

[56]  W R Strohl,et al.  Compilation and analysis of DNA sequences associated with apparent streptomycete promoters. , 1992, Nucleic acids research.

[57]  J. Suh,et al.  Effects of extracellular ATP on the physiology of Streptomyces coelicolor A3(2). , 2008, FEMS microbiology letters.

[58]  S. Horinouchi,et al.  Expression of the Streptomyces coelicolor A3(2) ptpA gene encoding a phosphotyrosine protein phosphatase leads to overproduction of secondary metabolites in S. lividans. , 1996, FEMS microbiology letters.

[59]  M. Bibb,et al.  Stationary‐phase production of the antibiotic actinorhodin in Streptomyces coelicolor A3(2) is transcriptionally regulated , 1993, Molecular microbiology.

[60]  C. Pei-lin,et al.  A pair of two-component regulatory genesecrA1/A2 inS. coelicolor , 2004 .

[61]  Eriko Takano,et al.  A complex role for the γ‐butyrolactone SCB1 in regulating antibiotic production in Streptomyces coelicolor A3(2) , 2001 .

[62]  J. Martín,et al.  Engineering of regulatory cascades and networks controlling antibiotic biosynthesis in Streptomyces. , 2010, Current opinion in microbiology.

[63]  J. Caballero,et al.  The act cluster contains regulatory and antibiotic export genes, direct targets for translational control by the bldA tRNA gene of streptomyces , 1991, Cell.

[64]  S. Kelly,et al.  Biosynthesis of the Sesquiterpene Antibiotic Albaflavenone in Streptomyces coelicolor A3(2)* , 2008, Journal of Biological Chemistry.

[65]  M. Buttner,et al.  DevA, a GntR-Like Transcriptional Regulator Required for Development in Streptomyces coelicolor , 2006, Journal of bacteriology.

[66]  R. Wagner,et al.  The Streptomyces coelicolor GlnR regulon: identification of new GlnR targets and evidence for a central role of GlnR in nitrogen metabolism in actinomycetes , 2008, Molecular microbiology.

[67]  A. Davidson,et al.  Ligand recognition by ActR, a TetR-like regulator of actinorhodin export. , 2008, Journal of molecular biology.

[68]  D. Sherman,et al.  Comparative genomic hybridizations reveal absence of large Streptomyces coelicolor genomic islands in Streptomyces lividans , 2007, BMC Genomics.

[69]  D. Hopwood,et al.  Physical and genetic characterisation of the gene cluster for the antibiotic actinorhodin in Streptomyces coelicolor A3(2). , 1986, Molecular & general genetics : MGG.

[70]  M. Osburne,et al.  Environmental DNA Fragment Conferring Early and Increased Sporulation and Antibiotic Production in Streptomyces Species , 2005, Applied and Environmental Microbiology.

[71]  Sarah Jordan,et al.  Identification of Three New Genes Involved in Morphogenesis and Antibiotic Production in Streptomyces coelicolor , 2003, Journal of bacteriology.

[72]  D. Hopwood,et al.  Physical and genetic characterisation of the gene cluster for the antibiotic actinorhodin inStreptomyces coelicolor A3(2) , 1986, Molecular and General Genetics MGG.