Tuberculosis (TB) is a leading cause of preventable deaths accounting for over two million deaths per year. Natural spread of multiple drug resistant (MDR) TB is a major threat to public health. CDC has classified MDR strains of M. tuberculosis (MTB) in Class C, within the list of organisms with potential use in bioterrorism. Discovery of novel anti-mycobacterial drug targets is critically needed to combat these threats, especially the MDR TB. The MTB genome is unusually rich in genes for lipid metabolism. It is becoming increasingly clear that lipid metabolism plays critical roles in TB. The critical steps, that are uniquely required for infection and survival of the pathogens in a dormant state for decades before the pathogens develops active TB when the host becomes immunodeficient, can be ideal targets for novel anti-TB drugs. We postulate that tgs/wes genes (TG synthase/wax ester synthase genes) and lip genes are involved in virulence and in the survival of the pathogen under dormant conditions. We will test this hypothesis. 1) Elucidate the biochemical functions of tgs/wes gene products. a) Characterize the enzymatic activities of the tgs/wes gene products expressed in E. coli. b) Determine the biochemical consequences of disrupting each tgs/wes gene on lipid metabolism. 2) Determine the consequence of tgs/wes gene disruption on host-pathogen interactions, a) Determine whether any molecular changes relevant to induction of TG synthesis can be detected as MTB reaches the hypoxia-induced nonreplicating state in culture b) Determine whether mutants have altered ability to grow in macrophages and trigger cytokine production, c) Determine the virulence, persistence, and the ability of the tgs/wes mutants to go into dormancy in mice and cause infection when the host is immunocompromised. 3) Elucidate the biochemical functions of the lip genes a) Express and characterize the TG hydrolase and thioesterase activities of lip gene products expressed in E. coli. b) Disrupt each lip gene and determine the biochemical consequences. 4) Determine the effect of lip disruptants on host-pathogen interaction, a) Determine the ability of the lip gene disruptants to survive the hypoxia-induced nonreplicating state in culture, b) Determine the effects of lip gene disruption on growth in macrophages and cytokine production. c) Determine the effect of lip gene disruption on virulence, persistence and the ability to undergo dormancy and reactivation in immunocompromised host. 5) Elucidate the biosynthetic mechanisms involved in the production of dimycocerosylphthiocerol (DIM), a known virulence factor. Identification of the unique steps in lipid metabolism critical for the disease will allow a search for novel drugs directed at these targets.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Special Emphasis Panel (ZRG1-BM-1 (04))
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Sizemore, Christine F
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University of Central Florida
Other Basic Sciences
Schools of Medicine
United States
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Daniel, Jaiyanth; Kapoor, Nidhi; Sirakova, Tatiana et al. (2016) The perilipin-like PPE15 protein in Mycobacterium tuberculosis is required for triacylglycerol accumulation under dormancy-inducing conditions. Mol Microbiol 101:784-94
Daniel, Jaiyanth; Sirakova, Tatiana; Kolattukudy, Pappachan (2014) An acyl-CoA synthetase in Mycobacterium tuberculosis involved in triacylglycerol accumulation during dormancy. PLoS One 9:e114877
Sirakova, Tatiana D; Deb, Chirajyoti; Daniel, Jaiyanth et al. (2012) Wax ester synthesis is required for Mycobacterium tuberculosis to enter in vitro dormancy. PLoS One 7:e51641
Daniel, Jaiyanth; Oh, Tae-Jin; Lee, Chang-Muk et al. (2007) AccD6, a member of the Fas II locus, is a functional carboxyltransferase subunit of the acyl-coenzyme A carboxylase in Mycobacterium tuberculosis. J Bacteriol 189:911-7
Lee, Kil-Soo; Dubey, Vinod S; Kolattukudy, Pappachan E et al. (2007) Diacyltrehalose of Mycobacterium tuberculosis inhibits lipopolysaccharide- and mycobacteria-induced proinflammatory cytokine production in human monocytic cells. FEMS Microbiol Lett 267:121-8
Sirakova, Tatiana D; Dubey, Vinod S; Deb, Chirajyoti et al. (2006) Identification of a diacylglycerol acyltransferase gene involved in accumulation of triacylglycerol in Mycobacterium tuberculosis under stress. Microbiology 152:2717-25
Cardona, P-J; Soto, C Y; Martin, C et al. (2006) Neutral-red reaction is related to virulence and cell wall methyl-branched lipids in Mycobacterium tuberculosis. Microbes Infect 8:183-90
Oh, Tae-Jin; Daniel, Jaiyanth; Kim, Hwa-Jung et al. (2006) Identification and characterization of Rv3281 as a novel subunit of a biotin-dependent acyl-CoA Carboxylase in Mycobacterium tuberculosis H37Rv. J Biol Chem 281:3899-908
Deb, Chirajyoti; Daniel, Jaiyanth; Sirakova, Tatiana D et al. (2006) A novel lipase belonging to the hormone-sensitive lipase family induced under starvation to utilize stored triacylglycerol in Mycobacterium tuberculosis. J Biol Chem 281:3866-75
Daniel, Jaiyanth; Deb, Chirajyoti; Dubey, Vinod S et al. (2004) Induction of a novel class of diacylglycerol acyltransferases and triacylglycerol accumulation in Mycobacterium tuberculosis as it goes into a dormancy-like state in culture. J Bacteriol 186:5017-30

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