A fundamental question in immunology is how the innate immune system detects pathogenic microbes. Host health depends critically upon the ability of the innate immune system to respond rapidly and selectively to pathogens, while avoiding inappropriate responses to self or harmless commensal microbes. Our goal is to describe how this rapid, sensitive, and accurate detection of pathogens is achieved at a molecular level. We have been particularly interested in a family of cytosolic pathogen detectors called inflammasomes. Inflammasomes comprise a family of multiprotein complexes that assemble in the cytosol in response to noxious and infectious stimuli. Inflammasomes initiate innate immune responses by activating the Caspase-1 protease. In general, it is poorly understood how inflammasomes recognize pathogens and assemble to activate Caspase-1. To address these questions, our studies have focused on a sub-family of inflammasomes called the Naip/Nlrc4 inflammasomes. Naip proteins (of which there are four in C57BL/6 mice) and Nlrc4 are members of a broader superfamily of proteins called NLRs (nucleotide- binding domain, leucine rich repeat-containing proteins). In the past funding period, we found that Naip5 mediates specific cytosolic detection of bacterial flagellin, whereas Naip2 mediates specific cytosolic detection of the inner rod protein from diverse bacterial type III secretion systems. We have also developed novel biochemical assays to dissect ligand binding and inflammasome oligomerization, and have used these novel assays to show that a key biochemical function of the Naips is to recognize ligands and induce downstream oligomerization of Nlrc4. In this renewal application, we describe three aims that will dissect ligand binding, oligomerization, and the downstream in vivo effector functions of the Naip/Nlrc4 inflammasomes.
Aim 1 seeks to determine how specific bacterial ligands are recognized by Naip proteins. This will be significant because it will represent the first analysis of ligand binding by any mammalian NLR protein.
Aim 2 seeks to dissect the biochemical and cellular mechanisms of inflammasome assembly, a critical but poorly understood process. Lastly, Aim 3 will investigate how inflammasomes initiate effector responses in vivo. In particular, we seek to establish a novel link between inflammasome activation and eicosanoid lipid mediator production in vivo. Eicosanoids are known to be critical mediators of inflammation, but their production has not previously been linked to inflammasomes. We show in vivo that inflammasome activation results in eicosanoid production and a severe vascular leakage syndrome that can be rapidly fatal. Taken together, our studies of the Naip/Nlrc4 inflammasomes will provide novel insights into ligand recognition, assembly and in vivo function of this critical family of cytosolic immune detectors.

Public Health Relevance

Infectious diseases remain a major cause of global mortality and morbidity. The design of novel immunotherapeutics, adjuvants and vaccines is predicated on a better understanding of how the innate immune system detects pathogens and initiates protective responses. Our proposal aims to fulfill this goal by dissection of the molecular mechanisms by which intracellular bacterial pathogens are sensed by the immune system.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
2R01AI075039-06A1
Application #
8502213
Study Section
Innate Immunity and Inflammation Study Section (III)
Program Officer
Palker, Thomas J
Project Start
2007-07-01
Project End
2018-06-30
Budget Start
2013-07-01
Budget End
2014-06-30
Support Year
6
Fiscal Year
2013
Total Cost
$359,021
Indirect Cost
$124,021
Name
University of California Berkeley
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
124726725
City
Berkeley
State
CA
Country
United States
Zip Code
94704
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Nichols, Randilea D; von Moltke, Jakob; Vance, Russell E (2017) NAIP/NLRC4 inflammasome activation in MRP8+ cells is sufficient to cause systemic inflammatory disease. Nat Commun 8:2209
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De Leon, Justin A; Qiu, Jiazhang; Nicolai, Christopher J et al. (2017) Positive and Negative Regulation of the Master Metabolic Regulator mTORC1 by Two Families of Legionella pneumophila Effectors. Cell Rep 21:2031-2038
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Rauch, Isabella; Tenthorey, Jeannette L; Nichols, Randilea D et al. (2016) NAIP proteins are required for cytosolic detection of specific bacterial ligands in vivo. J Exp Med 213:657-65
Chavarría-Smith, Joseph; Mitchell, Patrick S; Ho, Alvin M et al. (2016) Functional and Evolutionary Analyses Identify Proteolysis as a General Mechanism for NLRP1 Inflammasome Activation. PLoS Pathog 12:e1006052

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