Identification and Characterization of a New Regulator, TagR, for Environmental Stress Resistance Based on the DNA Methylome of Streptomyces roseosporus

This study established a novel strategy for screening regulators of environmental stress resistance based on the DNA methylome and discovered a new regulator, TagR. The TagR-WTA transporter regulatory pathway improved the resistance and antibiotic yield of strains and has the potential for wide application. Our research provides a new perspective on the optimization and reconstruction of industrial actinomycetes. ABSTRACT DNA methylation is a defense that microorganisms use against extreme environmental stress, and improving resistance against environmental stress is essential for industrial actinomycetes. However, research on strain optimization utilizing DNA methylation for breakthroughs is rare. Based on DNA methylome analysis and KEGG pathway assignment in Streptomyces roseosporus, we discovered an environmental stress resistance regulator, TagR. A series of in vivo and in vitro experiments identified TagR as a negative regulator, and it is the first reported regulator of the wall teichoic acid (WTA) ABC transport system. Further study showed that TagR had a positive self-regulatory loop and m4C methylation in the promoter improved its expression. The ΔtagR mutant exhibited better hyperosmotic resistance and higher decanoic acid tolerance than the wild type, which led to a 100% increase in the yield of daptomycin. Moreover, enhancing the expression of the WTA transporter resulted in better osmotic stress resistance in Streptomyces lividans TK24, indicating the potential for wide application of the TagR-WTA transporter regulatory pathway. This research confirmed the feasibility and effectiveness of mining regulators of environmental stress resistance based on the DNA methylome, characterized the mechanism of TagR, and improved the resistance and daptomycin yield of strains. Furthermore, this research provides a new perspective on the optimization of industrial actinomycetes. IMPORTANCE This study established a novel strategy for screening regulators of environmental stress resistance based on the DNA methylome and discovered a new regulator, TagR. The TagR-WTA transporter regulatory pathway improved the resistance and antibiotic yield of strains and has the potential for wide application. Our research provides a new perspective on the optimization and reconstruction of industrial actinomycetes.

[1]  R. Gromadka,et al.  Functional Characterization of TetR-like Transcriptional Regulator PA3973 from Pseudomonas aeruginosa , 2022, International journal of molecular sciences.

[2]  X. Mao,et al.  m4C DNA methylation regulates biosynthesis of daptomycin in Streptomyces roseosporus L30 , 2022, Synthetic and systems biotechnology.

[3]  G. Sutton,et al.  Horizontal transfer and evolution of wall teichoic acid gene cassettes in Bacillus subtilis , 2021, F1000Research.

[4]  Xue-Song Zhang,et al.  Discovering multiple types of DNA methylation from bacteria and microbiome using nanopore sequencing , 2021, Nature Methods.

[5]  A. H. Wang,et al.  Crystal structure of the N-terminal domain of TagH reveals a potential drug targeting site. , 2020, Biochemical and biophysical research communications.

[6]  M. Koffas,et al.  Wall teichoic acids: Physiology and applications. , 2020, FEMS microbiology reviews.

[7]  J. Coppee,et al.  4-Methylcytosine DNA modification is critical for global epigenetic regulation and virulence in the human pathogen Leptospira interrogans , 2020, Nucleic acids research.

[8]  Cong-Zhao Zhou,et al.  Cryo-electron Microscopy Structure and Transport Mechanism of a Wall Teichoic Acid ABC Transporter , 2020, mBio.

[9]  E. Cascales,et al.  Fur-Dam Regulatory Interplay at an Internal Promoter of the Enteroaggregative Escherichia coli Type VI Secretion sci1 Gene Cluster , 2020, Journal of bacteriology.

[10]  M. Loessner,et al.  Structure and function of Listeria teichoic acids and their implications , 2020, Molecular microbiology.

[11]  H. Tan,et al.  Important role of a LAL regulator StaR in the staurosporine biosynthesis and high-production of Streptomyces fradiae CGMCC 4.576 , 2019, Science China Life Sciences.

[12]  J. Casadesús,et al.  The bacterial epigenome , 2019, Nature Reviews Microbiology.

[13]  X. Mao,et al.  FadR1, a pathway-specific activator of fidaxomicin biosynthesis in Actinoplanes deccanensis Yp-1 , 2019, Applied Microbiology and Biotechnology.

[14]  J. Casadesús,et al.  Regulation of bistability in the std fimbrial operon of Salmonella enterica by DNA adenine methylation and transcription factors HdfR, StdE and StdF , 2019, Nucleic acids research.

[15]  P. Talaga,et al.  Glycosylation of Staphylococcus aureus cell wall teichoic acid is influenced by environmental conditions , 2019, Scientific Reports.

[16]  Ni Wan,et al.  13C-Metabolic Flux Analysis Reveals the Metabolic Flux Redistribution for Enhanced Production of Poly-γ-Glutamic Acid in dlt Over-Expressed Bacillus licheniformis , 2019, Front. Microbiol..

[17]  R. Kingsley,et al.  Evolution of Salmonella within Hosts , 2018, Trends in microbiology.

[18]  R. Alduina,et al.  The SCO1731 methyltransferase modulates actinorhodin production and morphological differentiation of Streptomyces coelicolor A3(2) , 2018, Scientific Reports.

[19]  C. Josenhans,et al.  The core genome m5C methyltransferase JHP1050 (M.Hpy99III) plays an important role in orchestrating gene expression in Helicobacter pylori , 2018, bioRxiv.

[20]  X. Mao,et al.  Transposon-based identification of a negative regulator for the antibiotic hyper-production in Streptomyces , 2018, Applied Microbiology and Biotechnology.

[21]  Yaoqi Zhou,et al.  A survey of Type III restriction-modification systems reveals numerous, novel epigenetic regulators controlling phase-variable regulons; phasevarions , 2018, Nucleic acids research.

[22]  R. Morgan,et al.  N4-cytosine DNA methylation regulates transcription and pathogenesis in Helicobacter pylori , 2018, Nucleic acids research.

[23]  S. Bentley,et al.  Phase-variable methylation and epigenetic regulation by type I restriction-modification systems. , 2017, FEMS microbiology reviews.

[24]  Long Liu,et al.  Microbial response to environmental stresses: from fundamental mechanisms to practical applications , 2017, Applied Microbiology and Biotechnology.

[25]  Lingqia Su,et al.  Enhanced extracellular production of recombinant proteins in Escherichia coli by co-expression with Bacillus cereus phospholipase C , 2017, Microbial Cell Factories.

[26]  Scott D. Taylor,et al.  The action mechanism of daptomycin. , 2016, Bioorganic & medicinal chemistry.

[27]  S. Yang,et al.  Improvement of daptomycin production via increased resistance to decanoic acid in Streptomyces roseosporus. , 2016, Journal of bioscience and bioengineering.

[28]  Long Liu,et al.  Metabolic engineering of acid resistance elements to improve acid resistance and propionic acid production of Propionibacterium jensenii , 2016, Biotechnology and bioengineering.

[29]  X. Mao,et al.  DepR1, a TetR Family Transcriptional Regulator, Positively Regulates Daptomycin Production in an Industrial Producer, Streptomyces roseosporus SW0702 , 2016, Applied and Environmental Microbiology.

[30]  J. Casadesús,et al.  OxyR-dependent formation of DNA methylation patterns in OpvABOFF and OpvABON cell lineages of Salmonella enterica , 2015, Nucleic acids research.

[31]  J. Imlay.  Diagnosing oxidative stress in bacteria: not as easy as you might think. , 2015, Current opinion in microbiology.

[32]  Hui Jiang,et al.  Generation of the natamycin analogs by gene engineering of natamycin biosynthetic genes in Streptomyces chattanoogensis L10. , 2015, Microbiological research.

[33]  X. Mao,et al.  Transcriptional regulation of the daptomycin gene cluster in Streptomyces roseosporus by an autoregulator, AtrA. , 2015, The Journal of biological chemistry.

[34]  M. Inui,et al.  Thermal and Solvent Stress Cross-Tolerance Conferred to Corynebacterium glutamicum by Adaptive Laboratory Evolution , 2015, Applied and Environmental Microbiology.

[35]  Z. Deng,et al.  Recognition and cleavage of 5-methylcytosine DNA by bacterial SRA-HNH proteins , 2015, Nucleic acids research.

[36]  Katy C. Kao,et al.  Evolved Osmotolerant Escherichia coli Mutants Frequently Exhibit Defective N-Acetylglucosamine Catabolism and Point Mutations in Cell Shape-Regulating Protein MreB , 2014, Applied and Environmental Microbiology.

[37]  E. Brown,et al.  Taking aim at wall teichoic acid synthesis: new biology and new leads for antibiotics , 2013, The Journal of Antibiotics.

[38]  B. Levin,et al.  Normal Mutation Rate Variants Arise in a Mutator (Mut S) Escherichia coli Population , 2013, PloS one.

[39]  S. Walker,et al.  Wall teichoic acids of gram-positive bacteria. , 2013, Annual review of microbiology.

[40]  Jianping Xie,et al.  The underling mechanism of bacterial TetR/AcrR family transcriptional repressors. , 2013, Cellular signalling.

[41]  J. Casadesús,et al.  Programmed Heterogeneity: Epigenetic Mechanisms in Bacteria , 2013, The Journal of Biological Chemistry.

[42]  Jianping Xie,et al.  Transcriptional analysis of the effect of exogenous decanoic acid stress on Streptomyces roseosporus , 2013, Microbial Cell Factories.

[43]  T. Roemer,et al.  Discovery of wall teichoic acid inhibitors as potential anti-MRSA β-lactam combination agents. , 2013, Chemistry & biology.

[44]  W. Wohlleben,et al.  Genetic Analysis of SCO2997, Encoding a TagF Homologue, Indicates a Role for Wall Teichoic Acids in Sporulation of Streptomyces coelicolor A3(2) , 2011, Journal of bacteriology.

[45]  V. Fedorenko,et al.  β-Glucuronidase as a Sensitive and Versatile Reporter in Actinomycetes , 2011, Applied and Environmental Microbiology.

[46]  S. Walker,et al.  ABC transporters required for export of wall teichoic acids do not discriminate between different main chain polymers. , 2011, ACS chemical biology.

[47]  T. Le,et al.  Structures of the TetR-like simocyclinone efflux pump repressor, SimR, and the mechanism of ligand-mediated derepression. , 2011, Journal of molecular biology.

[48]  Y. Shirakihara,et al.  Crystal structures of the multidrug binding repressor Corynebacteriumglutamicum CgmR in complex with inducers and with an operator. , 2010, Journal of molecular biology.

[49]  M. Davies,et al.  Phase variation controls expression of Salmonella lipopolysaccharide modification genes by a DNA methylation-dependent mechanism , 2010, Molecular microbiology.

[50]  A. Desbois,et al.  Antibacterial free fatty acids: activities, mechanisms of action and biotechnological potential , 2010, Applied Microbiology and Biotechnology.

[51]  O. Kuipers,et al.  Bistability, epigenetics, and bet-hedging in bacteria. , 2008, Annual review of microbiology.

[52]  C. Su,et al.  Characterization of the multidrug efflux regulator AcrR from Escherichia coli. , 2007, Biochemical and biophysical research communications.

[53]  J. Roth,et al.  Origin of mutations under selection: the adaptive mutation controversy. , 2006, Annual review of microbiology.

[54]  Vance G Fowler,et al.  Daptomycin versus standard therapy for bacteremia and endocarditis caused by Staphylococcus aureus. , 2006, The New England journal of medicine.

[55]  A. Peschel,et al.  Cation-Induced Transcriptional Regulation of the dlt Operon of Staphylococcus aureus , 2006, Journal of bacteriology.

[56]  S. Leibler,et al.  Phenotypic Diversity, Population Growth, and Information in Fluctuating Environments , 2005, Science.

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

[58]  N Beales,et al.  Adaptation of Microorganisms to Cold Temperatures, Weak Acid Preservatives, Low pH, and Osmotic Stress: A Review. , 2004, Comprehensive reviews in food science and food safety.

[59]  Francis C. Neuhaus,et al.  A Continuum of Anionic Charge: Structures and Functions of d-Alanyl-Teichoic Acids in Gram-Positive Bacteria , 2003, Microbiology and Molecular Biology Reviews.

[60]  S. Hohmann.  Osmotic Stress Signaling and Osmoadaptation in Yeasts , 2002, Microbiology and Molecular Biology Reviews.

[61]  W. Waldhäusl,et al.  Polyunsaturated Eicosapentaenoic Acid Displaces Proteins from Membrane Rafts by Altering Raft Lipid Composition* , 2001, The Journal of Biological Chemistry.

[62]  C. Harwood,et al.  D-Alanine substitution of teichoic acids as a modulator of protein folding and stability at the cytoplasmic membrane/cell wall interface of Bacillus subtilis. , 2000, The Journal of biological chemistry.

[63]  D. Karamata,et al.  The tagGH operon of Bacillus subtilis 168 encodes a two‐component ABC transporter involved in the metabolism of two wall teichoic acids , 1995, Molecular microbiology.

[64]  K. O'Brien,et al.  Plasmid cloning vectors for the conjugal transfer of DNA from Escherichia coli to Streptomyces spp. , 1992, Gene.

[65]  F. Huber,et al.  The formation of daptomycin by supplying decanoic acid to Streptomyces roseosporus cultures producing the antibiotic complex A21978C , 1988 .

[66]  D. Ellwood.  The wall content and composition of Bacillus substilis var. niger grown in a chemostat. , 1970, The Biochemical journal.

[67]  Mithila Rajagopal,et al.  Envelope Structures of Gram-Positive Bacteria. , 2017, Current topics in microbiology and immunology.

[68]  I. Poxton.  Teichoic Acids, Lipoteichoic Acids and Other Secondary Cell Wall and Membrane Polysaccharides of Gram-Positive Bacteria , 2015 .

[69]  Yeo Joon Yoon,et al.  Characterization of negative regulatory genes for the biosynthesis of rapamycin in Streptomyces rapamycinicus and its application for improved production , 2014, Journal of Industrial Microbiology & Biotechnology.