Project Summary Antisynthetase syndrome (ASSD) is a chronic autoimmune disorder characterized by myositis and interstitial lung disease (ILD). A key feature of ASSD is the presence of autoantibodies against aminoacyl-tRNA synthe- tases (aaRSs), among which histidyl-tRNA synthetase (HARS or HisRS) causing anti-Jo-1 antibodies is the most common. aaRSs are the enzymes that attach amino acids to their appropriate tRNA and are thus essential components of the intracellular translation machinery. Interestingly, some aaRSs are also secreted and function extracellularly to regulate inflammation and immune responses. However, the mechanisms by which aaRSs are released from cells, become targets of autoimmunity, and cause symptoms are not well understood. Importantly, tRNA, a key partner of aaRS, has not been considered at all in the context of ASSD. There is growing evidence suggesting the involvement of endosomal Toll-like receptor TLR7 and type I interferon (IFN-I) signaling in sus- taining and spreading inflammation in the autoimmune disease. The ligands that activate TLR7 are single strand RNAs (ssRNAs). Therefore, we hypothesize that tRNAs or tRNA fragments bound to aaRSs play critical roles in the pathogenesis of ASSD. We propose a two-year project to explore the hypothesis using ASSD-relevant cells and to generate a novel and clinically relevant animal model for mechanistic study and future therapeutic devel- opment. The results obtained from this study would uncover a previously unrecognized involvement of tRNAs in the pathogenesis of ASSD and suggest new targets for therapeutic development of the disease.