Saturday, September 7, 2024
9/7/2024
PROJECT SUMMARY Autophagy, the process by which cells dispose of cellular contents, has been increasingly appreciated as an important pathway in antiviral host defenses. Intriguingly, recent studies demonstrated that autophagy and the antiviral type I IFN response are intricately connected, and several molecules known to play key roles in IFN-mediated immunity are also important regulators of autophagy. Among these molecules with dual roles in autophagy and IFN-mediated immunity are several TRIM (tripartite motif) protein family members. However, whereas the molecular mechanisms by which TRIM proteins act as antiviral restriction factors or regulate IFN responses have been well characterized, our knowledge about the role of TRIM proteins in autophagy in response to viral infection is still rudimentary. The proposed study builds on a recent discovery by the Gack laboratory that identified TRIM23 as a critical regulator of autophagy-mediated host defense against a broad range of viruses. TRIM23 interacts with and activates the innate immune molecule TBK1, promoting TBK1-mediated phosphorylation of the selective autophagy receptor p62, which ultimately triggers viral clearance and host resistance. Mechanistically, the N-terminal RING E3 ligase of TRIM23 induces atypical non-degradative K27-linked auto-ubiquitination of the C-terminal ARF GTPase domain, which is unique to TRIM23. ARF ubiquitination results in enhanced activity of TRIM23 to hydrolyze GTP, activate TBK1, and mediate virus resistance. TRIM23 depletion or gene-targeting in various cell types showed that TRIM23-mediated autophagy confers antiviral activity against several viruses including HSV-1. Using molecular, biochemical, cell biological and structural approaches combined with virus infection studies, we will define in precise detail how TRIM23 mediates autophagy during viral infection. This study will yield insight into the mechanisms of TRIM23 activation by upstream regulators during viral infection (Aim 1). We will further determine the molecular details of how TRIM23 activates TBK1 and the role the enzymatic activities of TRIM23 – E3 ligase and GTPase – play in TBK1 activation. Finally, we will determine the physiological relevance of TRIM23 for antiviral host resistance and viral pathogenesis using in vitro cell culture systems and infection studies in TRIM23 knockout mice (Aim 2). Our studies will provide a molecular understanding of a key pathway in the autophagy-mediated host defense, which may guide the rational design of new antivirals.
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