The innate immune system recognizes microbial pathogens using germ line encoded receptors which recognize conserved structures made by microbes. For example, the "pattern recognition receptor" TLR4 recognizes lipopolysaccharides produced as part of the cell walls of Gram-negative bacteria. When professional phagocytic cells such as macrophages and dendritic cells internalize and degrade microbes, structures may be released. For example, bacterial cell wall peptidoglycan may be cleaved to release muramyl dipeptide, a ligand for the cytosolic pattern recognition receptor Nod2. In this project, we will stretch the definition of a pattern recognition receptor to include the host enzyme hexokinase, a central enzyme in cellular glucose metabolism. We have shown that upon degradation of peptidoglycan in phagosomes a component sugar, N-acetylglucosamine, is freed up and transported into the cytosol where it interacts with hexokinase causing release of the enzyme from the mitochondrial surface. Hexokinase release from mitochondria initiates a signaling process culminating in activation of caspase-1 in an "inflammasome" complex which is responsible for processing and release of the key pro-inflammatory cytokines IL-1? and IL-18 (among other effects). Thus hexokinase is the "receptor" that detects the presence of this microbe-derived sugar. Endogenously produced N-acetylglucosamine is not normally found free in the cytosol and thus does not engage hexokinase. In this project we will characterize this innate immune mechanism further in three aims.
In aim one we will characterize the cell biology of hexokinase signaling in macrophages and dendritic cells and determine how the process is regulated so as not to be toxic to the cells.
In aim two we will define the role of the pathway in host defense through characterizing the function of the host enzyme N-acetylglucosamine kinase as a negative regulator in vitro and in vivo in knockout mice.
In aim three well will explor how the Gram-positive peptidoglycan recycling system can be manipulated to influence signaling by this pathway in immune cells and alter microbial pathogenesis.

Public Health Relevance

Growing evidence suggests a strong relationship between inflammation and metabolic diseases such as obesity, type 2 diabetes, metabolic syndrome, and heart disease, although the ways in which inflammatory signaling and metabolic processes might be related are unclear. In this project, we are characterizing a new inflammatory signaling pathway that makes use of immune cells'basic metabolic enzymes to detect bacteria and trigger release of inflammatory mediators.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM085796-06
Application #
8442856
Study Section
Immunity and Host Defense Study Section (IHD)
Program Officer
Dunsmore, Sarah
Project Start
2008-04-01
Project End
2016-02-29
Budget Start
2013-03-01
Budget End
2014-02-28
Support Year
6
Fiscal Year
2013
Total Cost
$318,450
Indirect Cost
$125,450
Name
Cedars-Sinai Medical Center
Department
Type
DUNS #
075307785
City
Los Angeles
State
CA
Country
United States
Zip Code
90048
Wolf, Andrea J; Liu, George Y; Underhill, David M (2016) Inflammatory properties of antibiotic-treated bacteria. J Leukoc Biol :
Wolf, Andrea J; Reyes, Christopher N; Liang, Wenbin et al. (2016) Hexokinase Is an Innate Immune Receptor for the Detection of Bacterial Peptidoglycan. Cell 166:624-36
O'Rourke, J G; Bogdanik, L; Yáñez, A et al. (2016) C9orf72 is required for proper macrophage and microglial function in mice. Science 351:1324-9
Müller, Sabrina; Wolf, Andrea J; Iliev, Iliyan D et al. (2015) Poorly Cross-Linked Peptidoglycan in MRSA Due to mecA Induction Activates the Inflammasome and Exacerbates Immunopathology. Cell Host Microbe 18:604-12
Ma, Jun; Becker, Courtney; Reyes, Christopher et al. (2014) Cutting edge: FYCO1 recruitment to dectin-1 phagosomes is accelerated by light chain 3 protein and regulates phagosome maturation and reactive oxygen production. J Immunol 192:1356-60
Parsons, Matthew W; Li, Li; Wallace, Aaron M et al. (2014) Dectin-2 regulates the effector phase of house dust mite-elicited pulmonary inflammation independently from its role in sensitization. J Immunol 192:1361-71
Shimada, Kenichi; Crother, Timothy R; Karlin, Justin et al. (2012) Oxidized mitochondrial DNA activates the NLRP3 inflammasome during apoptosis. Immunity 36:401-14
Kaplan, Amber; Ma, Jun; Kyme, Pierre et al. (2012) Failure to induce IFN-ýý production during Staphylococcus aureus infection contributes to pathogenicity. J Immunol 189:4537-45
Ma, Jun; Becker, Courtney; Lowell, Clifford A et al. (2012) Dectin-1-triggered recruitment of light chain 3 protein to phagosomes facilitates major histocompatibility complex class II presentation of fungal-derived antigens. J Biol Chem 287:34149-56
Wolf, Andrea J; Arruda, Andrea; Reyes, Christopher N et al. (2011) Phagosomal degradation increases TLR access to bacterial ligands and enhances macrophage sensitivity to bacteria. J Immunol 187:6002-10

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