Abstract Sepsis is the 10th leading cause of death in the US. An unresolved systemic cytokine storm caused by bacterial infection is a hallmark of sepsis. The robust acute inflammatory response, through Toll Like Receptors (TLRs) and interleukin-1R like receptors (ILRs), trigger detrimental effects including multi-organ failure. Despite extensive research, therapies for sepsis focus on the use of antimicrobials that lead to multi-drug resistance. Hence, an unmet scientific need is to understand the molecular regulation of anti-inflammatory responses that diminish the severity of tissue injury. Single immunoglobulin interleukin-1-related receptor (SIGIRR), which is also known as Toll/IL-1 receptor 8, exhibits an anti-inflammatory effect against TLRs and ILRs signaling. Recently, IL-37, which is a suppressor of innate immunity, has been identified as the SIGIRR ligand. Both IL-37 and SIGIRR have been recognized as major therapeutic targets to lessen cytokine storm, however, very little is known regarding the molecular regulation of SIGIRR stability. Receptor degradation, a negative feedback regulation of receptor function, is a highly regulated process by post-translational modification, such as phosphorylation and ubiquitination. Ubiquitination is a molecular signal for protein degradation in either the proteasome or lysosome. De-ubiquitination, which is mediated by deubiquitinating enzymes (DUBs), tightly controls protein stability by removal of ubiquitin chains from target proteins. In our preliminary data, we discovered that (i) SIGIRR is poly-ubiquitinated and degraded in the proteasome in response to its ligand binding; (ii) Ubiquitin-specific protease (USP13), a member of DUBs, targets and stabilizes SIGIRR by hydrolyzing the ubiquitin chains from SIGIRR; (iii) glycogen synthase kinase 3β (GSK3β) phosphorylates SIGIRR and interrupts the association between SIGIRR and USP13, thereby reducing SIGIRR stability; (iv) USP13 increases survival rate in experimental sepsis. These observations have led to the following hypothesis: USP13 ameliorates cytokine storm and septic shock through deubiquitination and stabilization of the anti-inflammatory receptor, SIGIRR. To evaluate this hypothesis we will determine 1) molecular signature of USP13-promoted SIGIRR stability; 2) if GSK3β-induced disruption of USP13/SIGIRR interaction lessens anti-inflammatory effects of SIGIRR; 3) if stabilization of SIGIRR by USP13 mitigates endotoxin-induced pro-inflammatory responses. In summary, this application provides molecular mechanisms by which SIGIRR is degraded via phosphorylation-driven ubiquitination. Our preliminary data has uncovered two previously unrecognized post-translational modifications of SIGIRR: phosphorylation and ubiquitination. Two mediators were revealed: GSK3β, which phosphorylates SIGIRR; and USP13, which de- ubiquitinates SIGIRR. These studies will be the first to elucidate that phosphorylation of SIGIRR promotes its ubiquitination by disassoci...