Reverse Genetic Analysis of Antiviral Resistance Mechanisms

Scripps Research Institute, The, La Jolla CA

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Abstract

We do not yet fully understand how host cells recognize and respond to viral infections, and our broad goal is[unreadable] to1 clarify mechanisms of anti-viral innate immunity in mammals. We have previously studied microbial[unreadable] components recognized by Toll-like receptors (TLRs) and established mouse models lacking each TLR.[unreadable] These studies revealed that viruses are recognized by TLR-dependent and -independent mechanisms and[unreadable] that recognition leads to induction of type I interferons (IFNs), cytokines critical for antiviral host defense in[unreadable] mammals. Although TLR signaling pathways leading to IFN production have been extensively studied, the[unreadable] molecular mechanisms of TLR-independent viral detection have remained poorly understood. Recently, we[unreadable] have focused on a family of potential cytoplasmic viral detectors and their signaling intermediates. We also[unreadable] have reason to believe that IFN-independent anti-viral host defense mechanisms operate in mammals,[unreadable] although we do not know precisely what these mechanisms might entail. In the present proposal, we plan to[unreadable] utilize mouse reverse genetics (gene targeting) to generate and analyzing mutant mice lacking molecules that[unreadable] have a strong likelihood of participating in antiviral responses. Combining targeted mutations with one another[unreadable] and using classical methods of pathway analysis, we hope to understand the how the presence of viruses is[unreadable] sensed, how signals are initiated, and how they are transduced to yield a defensive response. First, we will[unreadable] continue our studies of potential intracellular viral detectors and their signaling molecules leading to the[unreadable] production of type I IFNs and proinflammatory cytokines. Second, we will search for new candidate molecules[unreadable] potentially involved in signaling pathways that lead from from cytoplasmic viral detectors to permit type I IFN[unreadable] production. Toward this end, we will use expression cloning and yeast two-hybrid screening. Third, we will try[unreadable] to understand and characterize novel anti-viral host defense mechanisms. To do so we will rely upon data from[unreadable] the forward genetic initiatives pursued in Project 1 and Project 2. We will generate mice with mutations in[unreadable] homologues of candidate molecules identified by Drosophila forward genetics conducted by the Strasbourg[unreadable] group. We will also target paralogues of molecules identified by mouse forward genetics conducted by the La[unreadable] Jolla group. By comparing the phenotypes of these mice with those bearing mutations affecting pathways[unreadable] serving the TLRs or cytoplasmic viral detectors, we will develop a comprehensive understanding of the host[unreadable] response. The successful attainment of such an understanding is enormously important, since viral infections[unreadable] are one of the most important causes of death worldwide. Potentially, the new concepts that we develop may[unreadable] be applied to the improvement of anti-viral vaccines and to the development of small molecules that enforce[unreadable] anti-viral responses through interaction with newly identified molecules.