Deletion of Two-Component Regulatory Systems Increases the Virulence of Mycobacterium tuberculosis

ABSTRACT Two-component regulatory signal transduction systems are widely distributed among bacteria and enable the organisms to make coordinated changes in gene expression in response to a variety of environmental stimuli. The genome sequence of Mycobacterium tuberculosis contains 11 complete two-component systems, four isolated homologous regulators, and three isolated homologous sensors. We have constructed defined mutations in six of these genes and measured virulence in a SCID mouse model. Mice infected with four of the mutants (deletions of devR, tcrXY, trcS, and kdpDE) died more rapidly than those infected with wild-type bacteria. The other two mutants (narL and Rv3220c) showed no change compared to the wild-type H37Rv strain. The most hypervirulent mutant (devRΔ) also grew more rapidly in the acute stage of infection in immunocompetent mice and in gamma interferon-activated macrophages. These results define a novel class of genes in this pathogen whose presence slows down its multiplication in vivo or increases its susceptibility to host killing mechanisms. Thus, M. tuberculosis actively maintains a balance between its own survival and that of the host.

[1]  G. Bancroft,et al.  Characterization of Auxotrophic Mutants ofMycobacterium tuberculosis and Their Potential as Vaccine Candidates , 2001, Infection and Immunity.

[2]  William R. Jacobs,et al.  Microbial Pathogenesis of Mycobacterium tuberculosis: Dawn of a Discipline , 2001, Cell.

[3]  V. Deretic,et al.  An Essential Two-Component Signal Transduction System in Mycobacterium tuberculosis , 2000, Journal of bacteriology.

[4]  M. Saier,et al.  Response regulators of bacterial signal transduction systems: Selective domain shuffling during evolution , 1995, Journal of Molecular Evolution.

[5]  G. Bancroft,et al.  Site-Directed Mutagenesis of the 19-Kilodalton Lipoprotein Antigen Reveals No Essential Role for the Protein in the Growth and Virulence of Mycobacterium intracellulare , 1998, Infection and Immunity.

[6]  W. Benjamin,et al.  Expression, Autoregulation, and DNA Binding Properties of the Mycobacterium tuberculosis TrcR Response Regulator , 2002, Journal of bacteriology.

[7]  S. Dhandayuthapani,et al.  Elements of signal transduction in Mycobacterium tuberculosis: in vitro phosphorylation and in vivo expression of the response regulator MtrA , 1996, Journal of bacteriology.

[8]  J. Belisle,et al.  Isolation of genomic DNA from mycobacteria. , 1998, Methods in molecular biology.

[9]  J. Tyagi,et al.  Identification and cloning of genes differentially expressed in the virulent strain of Mycobacterium tuberculosis. , 1993, Gene.

[10]  B. Bloom,et al.  Tuberculosis Pathogenesis, Protection, and Control , 1994 .

[11]  E. Groisman,et al.  Mg2+ as an Extracellular Signal: Environmental Regulation of Salmonella Virulence , 1996, Cell.

[12]  J. Mekalanos,et al.  Two-Component Signal Transduction and Its Role in the Expression of Bacterial Virulence Factors , 1995 .

[13]  K. Altendorf,et al.  KdpD and KdpE, proteins that control expression of the kdpABC operon, are members of the two-component sensor-effector class of regulators , 1992, Journal of bacteriology.

[14]  G. Kaplan,et al.  TNF-α Controls Intracellular Mycobacterial Growth by Both Inducible Nitric Oxide Synthase-Dependent and Inducible Nitric Oxide Synthase-Independent Pathways1 , 2001, The Journal of Immunology.

[15]  James C. Sacchettini,et al.  Persistence of Mycobacterium tuberculosis in macrophages and mice requires the glyoxylate shunt enzyme isocitrate lyase , 2000, Nature.

[16]  K. Shinozaki,et al.  Two-component systems in plant signal transduction. , 2000, Trends in plant science.

[17]  J. Hinds,et al.  Enhanced gene replacement in mycobacteria. , 1999, Microbiology.

[18]  J. Flynn,et al.  The Inducible Nitric Oxide Synthase Locus Confers Protection against Aerogenic Challenge of Both Clinical and Laboratory Strains of Mycobacterium tuberculosis in Mice , 2001, Infection and Immunity.

[19]  William R. Jacobs,et al.  Complex lipid determines tissue-specific replication of Mycobacterium tuberculosis in mice , 1999, Nature.

[20]  V. Mizrahi,et al.  Construction and Phenotypic Characterization of an Auxotrophic Mutant of Mycobacterium tuberculosis Defective in l-Arginine Biosynthesis , 2002, Infection and Immunity.

[21]  B. Gicquel,et al.  Analysis of the phthiocerol dimycocerosate locus of Mycobacterium tuberculosis. Evidence that this lipid is involved in the cell wall permeability barrier. , 2001, The Journal of biological chemistry.

[22]  J. Heitman,et al.  Cyclic AMP-Dependent Protein Kinase Controls Virulence of the Fungal Pathogen Cryptococcus neoformans , 2001, Molecular and Cellular Biology.

[23]  D. Sherman,et al.  Regulation of the Mycobacterium tuberculosis hypoxic response gene encoding alpha -crystallin. , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[24]  T. Gingeras,et al.  Comparing genomes within the species Mycobacterium tuberculosis. , 2001, Genome research.

[25]  Dirk Schnappinger,et al.  Regulation of the Mycobacterium tuberculosis hypoxic response gene encoding α-crystallin , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[26]  T. Parish,et al.  Use of a flexible cassette method to generate a double unmarked Mycobacterium tuberculosis tlyA plcABC mutant by gene replacement. , 2000, Microbiology.

[27]  E. Groisman,et al.  Regulation of Salmonella Virulence by Two-Component Regulatory Systems , 1995 .

[28]  G. Rook,et al.  Pathogenesis of Pulmonary Tuberculosis: an Interplay of Tissue-Damaging and Macrophage-Activating Immune Responses—Dual Mechanisms That Control Bacillary Multiplication , 1994 .

[29]  J. Hoch,et al.  Two-component and phosphorelay signal transduction. , 2000, Current opinion in microbiology.

[30]  G. Kaplan,et al.  Virulence of a Mycobacterium tuberculosis clinical isolate in mice is determined by failure to induce Th1 type immunity and is associated with induction of IFN-α/β , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[31]  J. Gowrishankar,et al.  trans-Acting Mutations in Loci Other than kdpDE That Affect kdp Operon Regulation inEscherichia coli: Effects of Cytoplasmic Thiol Oxidation Status and Nucleoid Protein H-NS on kdpExpression , 2001, Journal of bacteriology.

[32]  J. Hoch,et al.  Two-component signal transduction , 1995 .

[33]  W. Jacobs,et al.  Attenuation of and Protection Induced by a Leucine Auxotroph of Mycobacterium tuberculosis , 2000, Infection and Immunity.

[34]  M. Federle,et al.  A Response Regulator That Represses Transcription of Several Virulence Operons in the Group A Streptococcus , 1999, Journal of bacteriology.

[35]  I. Orme.  Virulence of recent notorious Mycobacterium tuberculosis isolates. , 1999, Tubercle and lung disease : the official journal of the International Union against Tuberculosis and Lung Disease.

[36]  C. Locht,et al.  Transient Requirement of the PrrA-PrrB Two-Component System for Early Intracellular Multiplication of Mycobacterium tuberculosis , 2002, Infection and Immunity.

[37]  P. Gounon,et al.  Attenuation of virulence by disruption of the Mycobacterium tuberculosis erp gene. , 1998, Science.

[38]  JoAnne L. Flynn,et al.  Fate of Mycobacterium tuberculosis within Murine Dendritic Cells , 2001, Infection and Immunity.

[39]  Peter M Woollard,et al.  Characterization of a Mycobacterium tuberculosis H37Rv transposon library reveals insertions in 351 ORFs and mutants with altered virulence. , 2002, Microbiology.

[40]  B. Gicquel,et al.  An essential role for phoP in Mycobacterium tuberculosis virulence , 2001, Molecular microbiology.

[41]  V. Deretic,et al.  Mycobacterium tuberculosis signal transduction system required for persistent infections , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[42]  B. Barrell,et al.  Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence , 1998, Nature.

[43]  David G. Russell,et al.  Mycobacterium tuberculosis: here today, and here tomorrow , 2001, Nature Reviews Molecular Cell Biology.

[44]  S. Dhandayuthapani,et al.  Gene expression in mycobacteria: transcriptional fusions based on xylE and analysis of the promoter region of the response regulator mtrA from Mycobacterium tuberculosis , 1994, Molecular microbiology.