The cGas/STING cytosolic DNA sensing pathway detects microbial DNA and plays a critical role in host defense. Growing evidence indicates that self-DNA that accumulates in the cytosol also engages cGAS to incite inflammation. In addition, a series of gain-of-function mutations that result in constitutive activation of STING have now been associated with a debilitating auto-inflammatory disease known as SAVI (STING-Associated- Vasculopathy with onset in Infancy). These patients suffer from severe vasculitic lesions and progressive interstitial lung disease (ILD), and frequently succumb to treatment-resistant ILD-associated with fibrosis. Similar to the lung disorders associated with other inflammatory or fibrotic lung diseases, very little is known about the pathogenic mechanisms in these patients. Thus, a better understanding of the mechanisms responsible for these conditions is needed in order to develop more effective therapies. We have recently developed a murine model for the most common SAVI mutation, STINGV154M (VM), and shown that these mice recapitulate the human disease by a variety of criteria, including the development of inflammatory/fibrotic lung disease. These VM mice provide a unique experimental tool for exploring the cell types and molecular mechanisms responsible for the initiation and progression of fibrotic lung disease. Rag1-deficiency completely rescues VM mice from both lung inflammation and lung fibrosis pointing to a critical role of the adaptive immunity in VM ILD. The role of STING has been focused on myeloid cells and VM mice have an expanded and activated myeloid/neutrophil compartment. However, radiation chimera studies have identified a key role for radioresistant cells in lung fibrosis as lethally irradiated VM mice reconstituted with WT bone marrow stem cells develop extremely severe lung disease even though these chimeras completely lack any VM-derived hematopoietic cells. Preliminary data implicate lymphatic endothelial cells (LECs) as initiators of BALT formation in VM mice. The goal of this application is to further explore the radioresistant innate immune cells that are directly or indirectly activated by the VM mutation and define the mechanisms by which these cells promote lung inflammation. Our approach will involve: (1) the in vivo analysis of a novel VM conditional KI line crossed to endothelial-restricted Cre deleter strains and other relevant controls, (2) scRNAseq comparison of stromal populations including lymphatic and vascular endothelial cells isolated from WT and VM mice; and (3) analysis of endothelial cells from VM and WT mice following in vitro activation by the VM mutation or more conventional STING agonists. There is an urgent need to identify better therapies for patients afflicted with autoimmune and autoinflammatory lung disorders and the studies proposed in this application should provide critical insights that will enable us to design the most relevant therapeutic targets. Further, these...