PROJECT SUMMARY Double-stranded RNA (dsRNA) is a well-known pathogen-associated molecular pattern (PAMP) generated during viral infections that triggers the innate immune response and plays a critical role in virus protection through the direct effect on type I IFN. However, in recent years it has been clear that this pathway can also be induced by endogenous dsRNA, with mitochondria transcripts being one of the major sources of endogenous dsRNA. The presence of endogenous dsRNA suggests that there must be mechanisms in place to keep on check their accumulation to avoid triggering RLRs pathways. Although there is a number of studies on how the production of type I IFN resulting from endogenous dsRNA can indirectly modulate T cell response, no previous studies have examined dsRNA in T cells. Our recent studies have revealed the presence of endogenous dsRNA in CDS cells upon activation, and mitochondria seems to be the primary source. In addition, we identify a new mechanism that CDS cells use to restrict the levels of dsRNA generated: inactivation of mitochondria GSK313 by phosphorylation on Ser389. Interestingly, failure to inactivate GSK313 by phospho-Ser389 results in a greater accumulation of mitochondrial dsRNA in activated CDS cells. Importantly, we also found that failure to inactivate GSK313 by phospho-Ser389 results in higher levels of IFNy produced by CDS cells. We hypothesize that mitochondrial dsRNAs are generated during activation of COB cells, can trigger the RLR pathway and contribute to sustain IFN production. We also propose that mitochondrial GSK3f3 interferes with the mitochondrial degradosome and that inactivation of mitochondrial GSK3/3 through phosphorylation on Ser389 in response to dsRNA is essential for maintaining dsRNA-homeostasis and restricting IFNr production. We will test this hypothesis with the following specific aims: 1) to show that mitochondrial dsRNA is generated during activation of CDS cells and contributes to the production of IFNy. 2) to show that inactivation of GSK3(3 through Ser389- phosphorylation plays a role in the homeostasis of mitochondrial dsRNA during activation of CDS cells. The results from the proposed studies could be a major breakthrough since they will show how endogenous dsRNA can contribute to sustain cytokine production in activated CDS cells, and its potential impact on autoimmune disease.