COORDINATION OF METABOLISM AND VIRULENCE DURING INFECTION
Romeo Tony
Ohio State Universitycity: Columbus country: United States (us)
Grant 1R01AI097116-01 from National Institute Of Allergy And Infectious Diseases
Abstract: Salmonella infections are a significant cause of morbidity and mortality in both developed and developing nations. There are over 2600 serovars of Salmonella that exhibit a variety of host ranges and disease manifestations, which include gastroenteritis, bacteremia, metastatic infections and paratyphoid and typhoid fevers. In all cases, infection requires adaptation of the bacterium to distinct conditions of a number of host compartments, mediated in large part by changes in the expression of virulence and metabolism genes. Understanding these adaptations during the infection cycle is important for developing new strategies for preventing and treating infection. Although knowledge is limited, regulation of the virulence and metabolism genes of this organism are intricately linked through genetic circuitry involving SirA (Salmonella invasion regulator) and CsrA (carbon storage regulator). SirA is a DNA binding transcription factor that is the response regulator of the BarA-SirA two component signal transduction system. CsrA is an RNA binding protein that regulates mRNA translation and stability. CsrA activity is regulated by small noncoding RNAs (CsrB, CsrC), which sequester the CsrA protein. In turn, the transcription of csrB and csrC is activated by SirA. Regulation by this system responds to substrates and end products of carbon metabolism, which vary within host compartments, leading to the hypothesis that the status of carbon availability in large part governs adaptive transitions during the infection cycle. In Aim 1, a combination of genomic, bioinformatic, molecular genetic and biochemical approaches will be used to define the SirA and CsrA regulons, and thereby greatly increase our understanding of the regulatory links between metabolism and virulence. In Aim 2, several complementary approaches will be used to determine precisely when and where genes of this system are expressed and active during Salmonella infection of mice. This information will be evaluated in context with the regulons (defined in Aim 1) as well as the environmental and metabolic conditions that are known to influence these regulators in vitro. SirA and CsrA orthologs are highly conserved throughout the gamma-proteobacteria and are important for disease transmission and/or virulence in every species in which they have been examined. Thus, an understanding of the conditions and stimuli affecting SirA and CsrA activities and the mechanisms by which these proteins coordinate virulence and metabolic gene expression may be applicable to a broad range of pathogens. Antibiotic therapies are relatively ineffective and are contraindicated for Salmonella gastrointestinal infections, which nevertheless have the potential to progress to life-threatening infections in infants and immunocompromised patients. Salmonella and many other species of bacteria utilize an ancient regulatory system to control metabolism and virulence. Determining the conditions that govern this regulatory system within the mammalian host, and determining the mechanisms and genetic circuitry by which this system functions may facilitate vaccine design and suggest novel targets and strategies for antibiotic and probiotic therapies
Keywords: Acetates; Affect; Antibiotic Therapy; Antibiotics; Bacteremia; Bacteria; base; Behavior; Biochemical; Bioinformatics; Carbon; cell motility; Cells; Complementary DNA; Data; Developed Countries; Developing Countries; Disease; disease transmission; DNA; DNA Binding; Environment; Escherichia; Exhibits; Formates; Future; Gammaproteobacteria; Gastroenteritis; gastrointestinal infection; Gel; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic; Genetic Transcription; Genetic Translation; Genomics; Glucose; Glycogen; Immunocompromised Host; In Vitro; in vivo; Infant; Infection; Intestines; Invaded; Klebsiella; Knowledge; Legionella; Life; Link; Location; mathematical model; Mediating; member; Metabolic; Metabolism; Methods; Microbial Biofilms; Modeling; Molecular; Molecular Genetics; Morbidity - disease rate; Mortality Vital Statistics; mRNA Stability; Mus; Names; novel; Nutrient; Organism; Orthologous Gene; Outcome; Paratyphoid Fever; pathogen; Pathogenesis; Pathway interactions; Physiological; Play; prevent; Probiotics; Process; Processed Genes; Promotor (Genetics); Protein Biosynthesis; Proteins; Proteobacteria; Pseudomonas; quorum sensing; Regulation; Regulator Genes; Regulon; Relative (related person); Reporter; Resistance; response; RNA; RNA Sequences; RNA-Binding Proteins; Role; Salmonella; Salmonella enterica; Salmonella infections; Seminal; Signal Transduction; Site; SPI1 gene; Staging; Stimulus; Stress; System; Systemic infection; Systems Biology; Testing; transcription factor; Type III Secretion System Pathway; Typhoid Fever; Untranslated RNA; Vaccine Design; Vibrio; Virulence; Volatile Fatty Acids; Work; Yersinia
Relevance: Antibiotic therapies are relatively ineffective and are contraindicated for Salmonella gastrointestinal infections, which nevertheless have the potential to progress to life-threatening infections in infants and immunocompromised patients. Salmonella and many other species of bacteria utilize an ancient regulatory system to control metabolism and virulence. Determining the conditions that govern this regulatory system within the mammalian host, and determining the mechanisms and genetic circuitry by which this system functions may facilitate vaccine design and suggest novel targets and strategies for antibiotic and probiotic therapies
Project start date: 2011-12-01
Project end date: 2016-11-30
Budget start date: 1-DEC-2011
Budget end date: 30-NOV-2012
1R01AI097116-01 (2012): $386875
Sponsored Links Excellgen http://Excellgen.com
Grants awarded to Romeo Tony
BIOFILM FORMATION AND DISPERSAL MECHANISMS
Romeo Tony, Professor
University Of Floridacity: Gainesville country: United States (us)
Grant 5R01GM066794-09 from National Institute Of General Medical Sciences
Abstract: In nature, bacteria grow predominantly within sessile, matrix-enclosed communities known as biofilms, rather than as unattached planktonic cells. Biofilms protect resident bacteria and complicate many chronic infections by preventing immune function, compromising antimicrobial therapy, and dispersing planktonic cells that spread infection to distant body sites. Our long-term goal is to obtain fundamental understanding of the interrelated structural, enzymatic, and regulatory elements required for biofilm formation and dispersal as a prerequisite for developing approaches to combat biofilm-related infections. While diverse structural components and regulatory stragtegies affect biofilm formation, we hypothesize that there are a few critical factors that are of importance in many species, which are best studied in model organisms. Furthermore, one such factor is the RNA-binding protein CsrA (RsmA), a global regulator that controls biofilm formation in many species. In Escherichia coli, CsrA represses biofilm formation, while it activates biofilm dispersal and motility. The most important role of CsrA in biofilm formation is to inhibit translation and stimulate decay of pgaABCD mRNA, which is needed for the production and transport of poly-beta-1,6-N-acetyl-D-glucosamine (PGA). This polysaccharide adhesin stabilizes biofilms of diverse species and promotes disease transmission and/or virulence in certain pathogens. Regulators of pgaABCD gene expression have profound effects on biofilm development, incuding NhaR, a transcriptional activator, and CsrD, a novel Csr-system component. Our preliminary studies reveal two additional important regulatory systems that control biofilm formation and PGA production without affecting pgaABCD gene expression. In the next phase of this project, we propose to 1) Elucidate two novel regulatory mechanisms of PGA polysaccharide production and biofilm formation. We will use in vivo and vitro polysaccharide synthesis and gene expression assays, transposon mutagenesis, and other molecular genetic approaches to accomplish this aim. 2) Delineate the functions of the pga genes and PGA itself. Effects of nonpolar deletions and site-directed mutations on polysaccharide polymerization, modification, localization, and chemical properties will be studied by biochemical and microscopic approaches. Properties of the PGA polysaccharide that determine its role as an adhesin will be examined. 3) The molecular genetic and biochemical mechanisms underlying biofilm dispersal will be examined. We will systematically determine the effects of CsrA induction (in preformed biofilm) on PGA levels, structure, and localization during the dispersal process. In the unlikely case that PGA is not involved in dispersal, we will examine possible roles of other envelope components. Bacterial biofilms are surface-associated, matrix-enclosed microbial communities that are prevalent throughout the biosphere. In a variety of infections, biofilm formation protects the bacteria against the immune system and complicates antibiotic therapies. Biofilm-based infections are noted for causing disease in conjunction with the use of catheters, prosthetics and other devices. The long-term goal of this study is to obtain fundamental information about the interrelated structural, enzymatic and regulatory elements that are required for biofilm formation and dispersal. The resulting information will bolster efforts to develop new strategies and therapeutic approaches to combat biofilm-related infections
Keywords: Acetylglucosamine; Adsorption; Affect; Anabolism; Animal Model; Antibiotic Therapy; antimicrobial; Bacteria; Bacterial Adhesins; base; Behavior; Binding (Molecular Function); Biochemical; Biological Assay; bis(3`, 5`)-cyclic diguanylic acid; Carbohydrates; Carbon; Catheters; cell envelope; cell motility; Cell surface; Cells; Charge; chemical property; Chronic; combat; Communities; Cyclic AMP; Cyclic Nucleotides; Deacetylase; Deacetylation; depolymerization; Detection; Development; Devices; dimer; Disease; disease transmission; Distant; DNA-Binding Proteins; Elements; env Gene Products; Escherichia coli; Event; Family; Fractionation; Gene Expression; Genes; genetic analysis; genetic regulatory protein; Genetic Transcription; Glucosamine; glycosyltransferase; Goals; Health; Homologous Gene; Hydrophobicity; Image; immune function; Immune system; Immunoblotting; Immunoelectron Microscopy; In Vitro; in vivo; Infection; Infection prevention; LacZ Genes; Lipoprotein (a); Mediating; Membrane; Membrane Proteins; Messenger RNA; Metabolic Pathway; Microbial Biofilms; microbial community; Microscopic; Modification; Molecular Genetics; Monovalent Cations; Mutagenesis; Mutation; Nature; novel; Nucleotides; Operon; Outcome; pathogen; periplasm; Phase; polymerization; Polysaccharides; Preparation; Process; Production; Property; Prosthesis; Protein Biosynthesis; Proteins; Regulatory Element; Repression; Research; response; RNA-Binding Proteins; Role; Screening procedure; Signal Transduction; Signaling Molecule; Site; Structure; Study models; Surface; System; Tertiary Protein Structure; Testing; theories; Therapeutic; Transcription Coactivator; Translations; Untranslated RNA; Virulence
Project start date: 2003-09-19
Project end date: 2012-03-31
Budget start date: 1-APR-2011
Budget end date: 31-MAR-2012
PFA/PA: PA-07-070
5R01GM066794-09 (2011): $296306