The long-term goal is to define the diversity of biochemical mechanisms used by bacterial pathogens to obtain lipid nutrients from the host. The knowledge derived from this research will add significantly to the understanding of pathogen physiology and answer the question of whether fatty acid synthesis inhibitors have therapeutic potential.
Aim #1 will define the role of a recently discovered enzyme system, fatty acid kinase, in the acquisition of host fatty acids by two different Gram-positive pathogens: Staphylococcus aureus and Streptococcus pneumoniae. Fatty acid kinase consists of two interacting proteins: a kinase domain protein (FakA) and a fatty acid binding protein (FakB). A single fakA is found in pathogens, whereas multiple fakB genes are common. Fatty acid kinase represents novel enzymology, and a thorough biochemical characterization of the FakA/B system will be performed to reveal FakB fatty acid binding selectivity, FakB as a fatty acid exchange protein, FabA-FakB interactions, and determine whether FakB-acyl-phosphate shuttles acyl-phosphate to membrane acyltransferases. X-ray crystallography and site-directed mutagenesis will define the mechanism of fatty acid kinase. The marked deficiency in virulence factor production exhibited by fakA and fakB strains shows fatty acid kinase also plays an important role in pathogenesis. An innovative, high impact idea proposes that transcriptional regulation of virulence factors by the FakA/B system occurs through the transfer of the phosphate on FakB-acyl-phosphate to the receiver domains of 2-component system response regulators to activate transcription. Establishing how fatty acid kinase modulates virulence factor transcription will define a potent, new regulatory circuit.
Aim #2 will test a new model for the origin of phospholipids required for the replication of the intracellular parasite Chlamydia trachomatis, and will determine importance of bacterial fatty acid synthesis to C. trachomatis physiology. C. trachomatis is an obligate intracellular bacterium that relies heavily on its cellulr host for nutrients. C. trachomatis has the enoyl-acyl carrier protein reductase of bacterial fatty acid synthesis, and a reductase-specific inhibitor will be used to establish the importance of de novo fatty acid biosynthesis in bacterial replication, and determine if enoyl reductase-targeted inhibitors could be deployed as therapeutics. The proposal that C. trachomatis produces phospholipids via a glycerol-phosphate acyltransferase that is not been previously reported in bacteria will be validated. Lipidomics, metabolic labeling and the biochemical characterization of the acyltransferases will critically test this new model for C. trachomatis lipid metabolism. Establishing that C. trachomatis triggers the host endoplasmic reticulum stress response will reveal an important pathway for the pathogen to enhance the supply of host phospholipids and optimize proliferation. Accomplishing these aims will define the importance of the acquisition of host lipids to the growth and virulence of three distinct groups of bacterial pathogens and serve as a blueprint for the understanding the physiology of other pathogens with similar lifestyles.

Public Health Relevance

The increasing prevalence of drug-resistant bacterial infections lends urgency to the development of effective new antibiotic countermeasures. The research will define a kinase pathway for the utilization of host fatty acids for membrane biogenesis in Gram-positive pathogens, and determine how it activates virulence factor production. The therapeutic potential of targeting fatty acid metabolism will be evaluated in several pathogens.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM034496-31
Application #
8814478
Study Section
Prokaryotic Cell and Molecular Biology Study Section (PCMB)
Program Officer
Chin, Jean
Project Start
1984-12-01
Project End
2018-11-30
Budget Start
2015-01-01
Budget End
2015-11-30
Support Year
31
Fiscal Year
2015
Total Cost
$729,800
Indirect Cost
$319,800
Name
St. Jude Children's Research Hospital
Department
Type
DUNS #
067717892
City
Memphis
State
TN
Country
United States
Zip Code
38105
Yao, Jiangwei; Rock, Charles O (2017) Bacterial fatty acid metabolism in modern antibiotic discovery. Biochim Biophys Acta 1862:1300-1309
Ericson, Megan E; Subramanian, Chitra; Frank, Matthew W et al. (2017) Role of Fatty Acid Kinase in Cellular Lipid Homeostasis and SaeRS-Dependent Virulence Factor Expression in Staphylococcus aureus. MBio 8:
Honsa, Erin S; Cooper, Vaughn S; Mhaissen, Mohammed N et al. (2017) RelA Mutant Enterococcus faecium with Multiantibiotic Tolerance Arising in an Immunocompromised Host. MBio 8:
Robertson, Rosanna M; Yao, Jiangwei; Gajewski, Stefan et al. (2017) A two-helix motif positions the lysophosphatidic acid acyltransferase active site for catalysis within the membrane bilayer. Nat Struct Mol Biol 24:666-671
Arensdorf, Angela M; Dillard, Miriam E; Menke, Jacob M et al. (2017) Sonic Hedgehog Activates Phospholipase A2 to Enhance Smoothened Ciliary Translocation. Cell Rep 19:2074-2087
Yao, Jiangwei; Rock, Charles O (2017) Exogenous fatty acid metabolism in bacteria. Biochimie 141:30-39
Yao, Jiangwei; Bruhn, David F; Frank, Matthew W et al. (2016) Activation of Exogenous Fatty Acids to Acyl-Acyl Carrier Protein Cannot Bypass FabI Inhibition in Neisseria. J Biol Chem 291:171-81
Subramanian, Chitra; Yun, Mi-Kyung; Yao, Jiangwei et al. (2016) Allosteric Regulation of Mammalian Pantothenate Kinase. J Biol Chem 291:22302-22314
Yao, Jiangwei; Rock, Charles O (2016) Resistance Mechanisms and the Future of Bacterial Enoyl-Acyl Carrier Protein Reductase (FabI) Antibiotics. Cold Spring Harb Perspect Med 6:a027045
Broussard, Tyler C; Miller, Darcie J; Jackson, Pamela et al. (2016) Biochemical Roles for Conserved Residues in the Bacterial Fatty Acid-binding Protein Family. J Biol Chem 291:6292-303

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