Molecular Regulation of MS Susceptibility Genes. Multiple sclerosis (MS) is an autoimmune disorder characterized by dysregulated T cell responses to antigenic components of myelin in the central nervous system (CNS). Although mechanisms leading to initiation of dysregulated T cell responses to myelin-derived antigens are poorly understood, discovery of a new subset of effector CD4+ T cells, Th17 cells, has transformed our understanding of MS disease pathogenesis and promises novel approaches to therapeutic intervention. In recent studies of the role of Th17 cells in CNS inflammation, we have discovered a novel mechanism by which production of the immunomodulatory cytokine, IL-10, is regulated. Specifically, we have identified an important transcriptional repressor that governs the production of IL-10 by CD4+ T cells - growth factor independent-1 (Gfi1). We find that in mice with T cell-specific deficiency of Gfi1 disease clinical disease and CNS inflammation are significantly ameliorated in an EAE model. Blockade of IL-10 signaling largely reverses protection conferred by Gfi1 deficiency. This strongly suggests that suppression of disease occurs via a mechanism linking Gfi1 to repression of IL10 gene expression in CD4+ T cells, and perhaps other immune cells that can produce IL-10. Importantly, in some individuals at increased risk for MS, the extended locus surrounding the GFI1 gene contains two single-nucleotide polymorphisms (SNPs), homozygosity for which leads to increased expression of GFI1 by activated peripheral blood cells. Further, GFI1 expression is elevated in peripheral blood of patients who later develop MS, consistent with the possibility that dysregulated GFI1 expression is an important risk factor for MS. While the data linking altered GFI1 expression to MS are only correlative at present and it is unknown how elevated expression of this factor might contribute to heightened susceptibility to MS, our preliminary studies have identified a mechanism that could explain a major component of Gfi1's linkage to MS and will be explored in this proposal. Specifically, we hypothesize that GFI1 is a major transcriptional repressor of IL10 gene expression and is central to IL10 gene regulation. Further, elevated expression of GFI1, as is found in association with MS susceptibility SNPs in the GFI1/EVI5 locus, confers impaired IL10 expression by immune cells that predisposes to, or exacerbates, CNS inflammation. To test this hypothesis, the role of Gfi1 in regulating human IL10 expression will be delineated in the context of CNS inflammation and we will define the function of cis-regulatory elements (CREs) in the IL10 locus that are targeted by Gfi1. These studies will employ a novel gene-targeting system that we have recently developed and implemented to enable site-specific, single copy integration of the human IL10 gene locus into the mouse genome. This represents a substantial technical advance with which to critically examine, for the first time, the function of distal CREs in chromatin remodeling and gene regulation. Using this model, we will be able to define the function of Gfi1-binding CREs in the human IL10 locus by examining the effects of mutation of these elements on IL10 gene remodeling, expression and protection in the EAE model.
Molecular Regulation of MS Susceptibility Genes. Multiple sclerosis (MS) is a disease of immune dysregulation mediated by CD4 T cells that have poorly restrained reactivity to components of the central nervous system. This proposal will address mechanisms by which cytokines control the maintenance and pathogenic potential of effector CD4+ T cells that contribute to MS pathogenesis. We expect that results from these studies will provide a basis for new interventions to curb pathogenic immunity to cellular components of the CNS that cause MS.