Vimentin-mediated regulation of the inflammasome in acute lung injury

Northwestern University At Chicago, Evanston IL

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Abstract

? DESCRIPTION (provided by applicant): The inflammasome is activated in response to a broad spectrum of infectious agents known to cause acute lung injury (ALI), but it must also be tightly regulated to avoid deleterious effects from the wide array of pathogens, pathogen-associated molecular patterns and danger-associated molecular patterns, which are known to trigger its activation. Activation of the inflammasome requires two distinct pro-inflammatory stimuli. In the first step, Toll-like receptor (TLR) signaling activates NF-?B and upregulates pro IL-1?. The second step involves the assembly of the multiprotein complex, which induces self-cleavage of caspase-1, and then mediates the cleavage of pro- IL-1? into its biologically active form. Growing evidence indicates that IL-1? is a critical mediator of acute inflammation and acute lung injury. We will test our hypothesis using two well- established murine models of NLRP3 and NLRC4 inflammasome activation, LPS and Pseudomonas aeruginosa, respectively. We will conduct experiments to demonstrate that NLR proteins, NLRP3 and NLRC4, via their leucine-rich-repeat (LRR) interact with vimentin and that this protein-protein interaction is required for the processing and maturation of pro-IL-1? into biologically active IL-1?. We will further examine whether the localization of the NLR proteins at the outer mitochondrial membrane is required for the interaction with vimentin during inflammasome activation. Our data suggest that the vimentin IF network acts as an additional checkpoint to control the activation of the inflammasome, a hypothesis we will test using a pharmacologic inhibitor of vimentin/NLR protein interactions in vitro and in vivo. These findings are likely to have important biological and clinical significance as targeting the interaction between NLRs and vimentin could provide a suite of novel therapeutics to treat patients with ALI/ARDS. We have formulated three interrelated specific aims to study the regulation of vimentin intermediate filaments in both in vivo and in vitro models of acute lung injury: Specific Aim 1: To determine the mechanism by which vimentin, a type III intermediate filament protein, activates the inflammasome, which is required for inflammation and acute lung injury. Specific Aim 2: To identify the protein:protein interactions between vimentin and NLRs, NLRP3 and NLRC4, required for the activation of the inflammasome. Specific Aim 3: To determine whether translocation of NLR proteins to the mitochondrial membrane is required for the vimentin-mediated assembly and activation of the inflammasome.