A5030322816- Tuberculosis Research and Epidemiology
- Mycobacterium research and diagnosis
- RNA and protein synthesis mechanisms
- HIV Research and Treatment
- HIV/AIDS drug development and treatment
- Bacterial Genetics and Biotechnology
- Pneumocystis jirovecii pneumonia detection and treatment
- HIV/AIDS Research and Interventions
- Bacteriophages and microbial interactions
- Immunodeficiency and Autoimmune Disorders
- Antibiotic Resistance in Bacteria
- Biochemical and Molecular Research
- Cytomegalovirus and herpesvirus research
- Microbial Natural Products and Biosynthesis
- RNA modifications and cancer
- Cancer therapeutics and mechanisms
- Pneumonia and Respiratory Infections
- Immune responses and vaccinations
- Enzyme Structure and Function
- Diagnosis and treatment of tuberculosis
- Infectious Diseases and Tuberculosis
- Viral Infections and Immunology Research
- Pharmacological Effects of Natural Compounds
- Microbial Metabolism and Applications
- Streptococcal Infections and Treatments
Harvard University
2014-2024
Boston Children's Hospital
2014-2024
Harvard University Press
1987-2005
Boston Children's Museum
2005
In-Q-Tel
2003
Yonsei University
2003
Brigham and Women's Hospital
1999
Duquesne University
1999
National Cancer Institute
1990-1994
National Institutes of Health
1990-1993
The Mycobacterium tuberculosis genome contains 11 serine/threonine kinase genes including two, pknA and pknB , that are part of an operon encoding involved in cell shape control wall synthesis. Here we demonstrate predominantly expressed during exponential growth, overexpression these kinases slows growth alters morphology. We determined the preferred substrate motifs PknA PknB, identified three vivo substrates kinases: PknB; Wag31, ortholog division protein DivIVA; Rv1422, a conserved...
The Mycobacterium tuberculosis genome encodes 11 serine/threonine protein kinases (STPKs) that are structurally related to eukaryotic kinases. To gain insight into the role of Ser/Thr phosphorylation in this major global pathogen, we used a phosphoproteomic approach carry out an extensive analysis M. . We identified more than 500 events 301 proteins involved broad range functions. Bioinformatic quantitative vitro kinase assays on peptides containing subset these sites revealed dominant motif...
mariner family transposons are widespread among eukaryotic organisms. These apparently horizontally transmitted diverse eukaryotes and can also transpose in vitro the absence of added cofactors. Here we show that derived from element Himar1 efficiently bacteria vivo . We have developed simple transposition systems by using minitransposons, made up short inverted repeats flanking antibiotic resistance markers. elements after expression transposase an appropriate bacterial promoter. found...
ABSTRACT Mycobacterium tuberculosis is a specialized intracellular pathogen that must regulate gene expression to overcome stresses produced by host defenses during infection. SigH an alternative sigma factor we have previously shown plays role in the response stress of saprophyte smegmatis . In this work investigated sigH M heat and oxidative stress. We determined mutant more susceptible inducible thioredoxin reductase/thioredoxin genes trxB2/trxC unknown function, Rv2466c, regulated via...
The Mycobacterium tuberculosis genome contains 11 serine/threonine kinase genes, and the products of two these, PknA PknB, are key components a signal transduction pathway that regulates cell division and/or morphology. Previously, we have shown one substrate these kinases is Wag31, homologue protein DivIVA present, but not known to be phosphorylated, in other Gram-positive bacteria. Here, investigate localization function Wag31 its phosphorylation. We demonstrate localized poles. further...
2′,3′-Dideoxyinosine (ddI) is a dideoxynucleoside with potent activity in vitro against the human immunodeficiency virus (HIV). In initial clinical trials adults, ddI showed evidence of antiretroviral little hematologic toxicity.
A system that permits molecular genetic manipulation of mycobacteria was developed on the basis yeast paradigm gene replacement by homologous recombination. shuttle vector can replicate autonomously at a high copy number in Escherichia coli but must integrate into DNA for survival Mycobacterium smegmatis constructed. The contains ColE1 origin replication, antibiotic resistance markers ampicillin and kanamycin, nutritional marker (pyrF) allows both positive negative selection E. M. smegmatis,...
Despite many methodological advances that have facilitated investigation of Mycobacterium tuberculosis pathogenesis, analysis essential gene function in this slow-growing pathogen remains difficult. Here, we describe an optimized CRISPR-based method to inhibit expression genes based on the inducible enzymatically inactive Cas9 protein together with gene-specific guide RNAs (CRISPR interference). Using system target several M. tuberculosis, achieved marked inhibition resulting growth...
The Mycobacterium tuberculosis Ser/Thr kinase PknB has been implicated in the regulation of cell growth and morphology this organism. extracytoplasmic domain membrane protein comprises four penicillin binding associated (PASTA) domains, which are predicted to bind stem peptides peptidoglycan. Using a comprehensive library synthetic muropeptides, we demonstrate that binds muropeptides manner dependent on presence specific amino acids at second third positions peptide, sugar moiety...
The Mycobacterium tuberculosis genome contains an unusually high number of toxin–antitoxin modules, some which have been suggested to play a role in the establishment and maintenance latent tuberculosis. Nine these loci belong mazEF family, encoding intracellular toxin MazF its antitoxin inhibitor MazE. Nearly every ortholog recognizes unique three- or five-base RNA sequence cleaves mRNA. As result, toxins selectively target subset transcriptome for degradation are known as “mRNA...
Toxin-antitoxin (TA) systems play key roles in bacterial persistence, biofilm formation and stress responses. The MazF toxin from the Escherichia colimazEF TA system is a sequence- single-strand-specific endoribonuclease, many studies have led to proposal that family members exclusively target mRNA. However, recent data indicate some toxins can cleave specific sites within rRNA concert with In this report, we identified repertoire of RNAs cleaved by Mycobacterium tuberculosis MazF-mt9 using...
Bacille Calmette-Guérin (BCG), the live attenuated tuberculosis vaccine, is manufactured under different conditions across globe generating formulations that may differ in clinical efficacy. Innate immune recognition of BCG contributes to immunogenicity suggesting differences viability contribute divergent activity licensed formulations.We compared BCG-Denmark (DEN), -Japan (JPN), -India (IND), -Bulgaria (BUL) and -USA vitro with respect a) as measured by colony-forming units (CFU),...
Human immunodeficiency virus type 1 (HIV-1) strains were isolated from nine patients before and after prolonged therapy with either an alternating regimen of 3'-azido-3'-deoxythymidine (AZT) 2',3'-dideoxycytidine (ddC) (AZT/ddC) or 2',3'-dideoxyinosine (ddI) alone. All obtained four who received AZT/ddC for up to 41 mo highly insensitive AZT in vitro. Only one strain showed reduced susceptibility ddC addition had previously unreported amino acid substitutions the viral polymerase-encoding...
ABSTRACT Mycobacterium tuberculosis , an obligate mammalian pathogen, adapts to its host during the course of infection via regulation gene expression. Of regulators transcription that play a role in this response, several alternative sigma factors M. have been shown control expression response stresses, and some these are required for virulence or persistence vivo. For study, we examined factor SigD virulence. Using microarray analysis, identified genes whose was altered strain with sigD...
Summary mRNA interferases are sequence‐specific endoribonucleases encoded by toxin‐antitoxin (TA) systems in bacterial genomes. Previously, we demonstrated that Mycobacterium tuberculosis contains at least seven genes encoding MazF homologues (MazF‐mt1 to ‐mt7) and determined cleavage specificities for MazF‐mt1 MazF‐mt6. Here have developed a new general method the determination of recognition sequences longer than three bases with use phage MS2 RNA as substrate CspA, an chaperone, which...
Toxin–antitoxin (TA) systems are implicated in the downregulation of bacterial cell growth associated with stress survival and latent tuberculosis infection, yet activities intracellular targets these TA toxins largely uncharacterized. Here, we use a specialized RNA-seq approach to identify Mycobacterium VapC toxin, VapC-mt4 (also known as VapC4), which have eluded detection using conventional approaches. Distinct from one other characterized toxin M. that cuts 23S rRNA at sarcin–ricin loop,...
Tuberculosis is the leading killer among infectious diseases worldwide. Increasing multidrug resistance has prompted new approaches for tuberculosis drug development, including targeted inhibition of virulence determinants and signaling cascades that control many downstream pathways. We used a multisystem approach to determine effects potent small-molecule inhibitor essential
Summary SigH, an alternative sigma factor of Mycobacterium tuberculosis , is a central regulator the response to oxidative and heat stress. Exposure these stresses results in increased expression sigH itself, genes encoding additional regulators effectors bacterial stresses. In this work we show that RshA, protein encoded by gene operon, anti‐sigma SigH. We demonstrate RshA binds SigH vitro vivo . This protein–protein interaction, as well ability inhibit SigH‐dependent transcription,...
ABSTRACT Extracytoplasmic function (ECF) sigma factors are a heterogeneous group of alternative that regulate gene expression in response to variety conditions, including stress. We previously characterized mycobacterial ECF factor, SigE, contributes survival following several distinct stresses. A encoding closely related sigH , was cloned from Mycobacterium tuberculosis and smegmatis . single copy this is present these other fast- slow-growing mycobacteria, M. fortuitum avium While the...
The extracytoplasmic function (ECF) sigma factors constitute a diverse group of alternative that have been demonstrated to regulate gene expression in response environmental conditions several bacterial species. Genes encoding an ECF factor Mycobacterium tuberculosis, avium, and smegmatis, designated sigE, were cloned analyzed. Southern blot analysis the presence single copy this these species bovis BCG, leprae, fortuitum. Sequence showed sigE be highly conserved among M. leprae. Recombinant...
Human T cells reactive to Mycobacterium tuberculosis were cloned from peripheral blood mononuclear (PBMC) of four patients by using whole irradiated bacilli as the in vitro stimulatory agent. Twenty-two cell clones (CD4+) tested for their reactivity 12 different mycobacterial species and showed a distribution limited broad cross-reactivity. These also three abundant M. proteins 65, 19, 14 kD, each expressed recombinant DNA Escherichia coli. The that previously isolated lambda gt11 genomic...
Mycobacterium tuberculosis sigL encodes an extracytoplasmic function (ECF) sigma factor and is adjacent to a gene for membrane protein (Rv0736) that contains conserved HXXXCXXC sequence. This motif found in anti-sigma factors regulate several ECF factors, including those control oxidative stress responses. In this work, SigL Rv0736 were be cotranscribed, the intracellular domain of was shown interact specifically with SigL, suggesting may encode SigL. An M. mutant not more susceptible than...