Project Summary Multiple Sclerosis (MS) is an inflammatory, demyelinating disease of the central nervous system (CNS). Increasing evidence indicates that the inhibition of remyelination is a major cause of the permanent neurodegeneration. However the precise mechanism that inhibits OPCs differentiation and remyelination remains an active area of research. While inflammation-associated NOTCH1 activation was implicated in defective remyelination in MS, the precise mechanism by which NOTCH1 was engaged during inflammatory response remains elusive. Using experimental autoimmune encephalomyelitis (EAE), an animal model of MS, we and others have established that IL-17 signaling is crucial for the induction of inflammation in the CNS. We have now discovered a direct integration of IL-17 and NOTCH1 signaling that plays a dominant role in impairing the differentiation of OPCs. IL-17 stimulation induced the release of the intracellular domain of NOTCH1 (NICD1) in the OPCs co-cultured with astrocytes. Mechanistically, IL-17R interacts with NOTCH1 via the extracellular domain, which facilitates the cleavage of NICD1. Subsequently, Act1, the adaptor protein for IL-17 signaling, forms a complex with NICD1, followed by translocation of Act1-NICD1 complex into the nucleus. Act1-NICD1 complex promotes the assembly of RBP-J containing transcriptional complex on the promoters of NOTCH1 target genes implicated in CNS inflammation. Furthermore, a decoy peptide disrupting the IL-17RA–NOTCH1 interaction inhibited IL-17-induced NICD cleavage, reduced IL-17-induced OPC prolifereation and attenuated the myelin loss in EAE model. Based on these findings, we hypothesize IL-17-NOTCH1 pathway drives the expression of a specific set of genes to promote inflammation and inhibition of OPC differentiation, thereby impairing the remyelination process in the demyelinating disease. We will test this hypothesis through Aim 1: Investigate the molecular mechanism of the inhibitory role of IL-17-NOTCH1 integration on OPC differentiation. Aim 2: Investigate the in vivo impact of IL-17-NOTCH1 integration on remyelination process. The completion of this proposal will provide fundamental insight into the critical integration of signaling pathways of IL-17 and NOTCH1, which underlies the pathogenesis of demyelinating disease, offering novel therapeutic strategies for MS patients by promoting remyelination.