Project Summary/Abstract Multiple sclerosis (MS) is both an inflammatory and neurodegenerative disease that has genetic and environmental components to susceptibility. The identification of susceptibility loci by unbiased genome-wide association studies (GWAS) is a major step forward in multiple sclerosis (MS), but we do not yet understand how allelic variation influences immune function. This transition remains a major challenge in the understanding of MS and of other human autoimmune diseases. Increasing evidence suggests that autoreactive CD4+ T cells play a central role in MS pathophysiology. Our evaluation of the 110 validated MS susceptibility loci using a cohort of healthy genotyped adults identified MS variants that alter nearby gene expression in naïve CD4+ T cells. We quickly focused our attention on the rs6908428 single nucleotide polymorphism (SNP) located near the transcription start site of the Abelson helper Integration site 1 (AHI1) gene because (i) this locus is robustly associated with MS susceptibility (p=1.8 x 10-20), (ii) the “credible set” of SNPs in strong linkage disequilibrium (LD) with the top SNP that may contain the causal variant is small (n=11), (iii) the susceptibility allele rs6908428A has a strong effect on RNA expression of AHI1 (1.12 x 10-107) but not on neighboring genes in CD4+ T cells (not a coincidental overlap), (iv) AHI1 expression is induced in pathogenic interleukin (IL)-17A-producing T cells (Th17) that are implicated in MS pathogenesis and are well characterized in the laboratory and (v) we have discovered that Ahi1/AHI1 binds phospholipase C (PLC)γ1 and that pharmacological inhibition of PLCγ1 suppresses human Th17 cell proliferation and a mouse model of MS. Finally, in MS patients, AHI1 RNA expression is elevated in CD4+ T cells from MS patients compared to healthy subjects, as well as in PBMCs of MS patients with a higher disease activity. The principal goals of the proposed project are: (1) Identification as well as in vitro and ex vivo validation of causal variant(s) involved in the regulation of AHI1 transcription in CD4+ T cells, (2) analysis of Ahi1 function in murine CD4+ T cells and its influence on encephalitogenicity in a mouse model of MS, and (3) analysis of AHI1's influence on gene networks in human Th17 cells in genotyped healthy subjects (n=100), and glatiramer acetate-treated MS patients (n=100). We thus propose a multifaceted approach integrating human genetic and human functional immunology experiments to understand how the susceptibility loci alter the state of activation in pathogenic Th17 to modulate susceptibility to MS and aggravate disease severity.