Elucidating the Role of cGAS-STING in Lung Tissue Repair and Remodeling

PROJECT SUMMARY Idiopathic pulmonary fibrosis is a chronic, progressive, highly heterogeneous fibrotic lung disease. The median survival from the time of diagnosis is ~3 years, demonstrating that IPF is considered more lethal than many cancers. There is no identifiable cause for most patients with IPF. Current FDA-approved drugs only slow progression but do not cure IPF. The pathophysiology of pulmonary fibrosis relies on repeated local micro- injuries to the alveolar epithelium triggering DNA damage, cell death, and aberrant lung tissue remodeling. Elevated levels of cell-free double-stranded DNA (dsDNA) serve as a negative predictive factor for fatal outcomes in IPF patients. dsDNA in the cytoplasm is recognized by cGAS-STING (stimulator of interferon genes); cGAS-STING is activated in fibrotic lung tissue from interstitial lung disease patients. cGAS-STING promotes the translocation of IFN regulatory factor 3 (IRF3) and NF-κB to the nucleus, inducing type I IFNs and other proinflammatory cytokines. Using a well-defined, murine model of pulmonary fibrosis, we find increased dsDNA in BALF of injured mice. Critically, Cgas-/- mice show reduced inflammation, decreased lung collagen levels, and improved static lung compliance compared to WT mice. Recently, STING was found to interact with vimentin, a protein critical for cell structure and organelle positioning. Given our previous report that Vimentin-/- mice are protected from bleomycin-induced lung injury and fibrosis, together with evidence that vimentin facilitates STING ER-to-Golgi trafficking, an essential and rate-limiting step in activating cGAS-STING, we hypothesize persistent activation of cGAS-STING signaling in alveolar epithelial cells and alveolar macrophages contributes to aberrant lung tissue repair. Specific Aim 1: Determine whether cGAS-STING-mediated IRF3 and NF-kB activation in monocyte- derived alveolar macrophages is necessary for aberrant lung tissue remodeling associated with pulmonary fibrosis. We will genetically inhibit cGAS-STING-mediated IRF3 and NF-kB signaling in MoAM during the fibrotic phase of bleomycin injury and use complementary physiologic and molecular techniques to assess lung repair. Specific Aim 2. Determine the mechanism by which VIM-cGAS-STING axis contributes to aberrant lung tissue repair and pulmonary fibrosis. We will define the mechanism by which VIM, a STING- interacting protein, traffics STING from the ER-to-Golgi, initiating STING-dependent IRF3 and NF-κB signaling leading to the expression of type I IFNs and inflammatory cytokines. Specific Aim 3. Determine whether cGAS- STING–PERK signaling is required for alveolar epithelial cell damage/stress and pulmonary fibrosis. Our data shows that cGAS-STING is activated in alveolar epithelial cells from patients with pulmonary fibrosis. Upon binding to cGAMP, STING binds to and directly activates the ER-located kinase PERK in the endoplasmic reticulum (ER). We will address whether the STING-PERK axis is required for alveolar epithelial cell damage/stress and contributes to aberrant lung tissue remodeling associated with pulmonary fibrosis.