The long term goal of this project is identification of the chromosomal locations and functions of genes controlling diabetes pathogenesis in NOD/Lt mice. Insulin dependent diabetes in NOD/Lt mice is under polygenic control and reflects development of a dysregulated immune system in which autoreactive T cells are neither deleted intrathymically nor suppressed peripherally. Genes within the major histocompatibility complex (MHC) are important components of the genetic susceptibility; however, non-MHC susceptibility genes interact with the diabetogenic MHC loci to initiate diabetes pathogenesis. Genetic analysis will be centered on NOR/Lt, a NOD- related, diabetes-free stock. This stock contains NOD alleles at MHC and most other non-MHC loci typed, but contains segments of 4 chromosomes derived from the C57BL/KsJ strain. Diabetes resistance in the NOR stock is accompanied by a reconstituted syngeneic mixed lymphocyte reaction (SMLR), a peripheral T cell response absent in NOD/Lt. Thus, presence or absence of SMLR activity serves as a valuable immunophenotypic marker to segregate analysis the C57BL/KsJ genes in NOR/Lt abrogating diabetes pathogenesis and relate them to immunophenotypic markers. This will be accomplished by development of NOD stocks congenic for chromosomal segments carrying diabetes resistance alleles from NOR.
The second aim i s to analyze by genetic segregation how pathogenesis-predisposing MHC haplotypes interact with diabetogenic non-MHC genes (defined in the first aim) to mediate pathogenesis via perturbation of immunoregulatory systems.
The third aim i s to develop monoclonal antibodies to permit characterization of Mrt-6, a mouse gene we mapped to Chr 7 and homologous to RT6 in the rat. RT6 is a T cell surface differentiation antigen defined in rats; a subset of RT6+ T cells possess immunoregulatory function since their depletion elicits diabetes in heretofore resistant rats. Northern blot analysis reveals lower Mrt-6 RNA expression in NOD compared to diabetes-resistant mice, indicating that Mrt-6 may also serve as a marker for immunoregulatory subsets in the NOD mouse.
The fourth aim i s to investigate the potential pathogenic significance of pancreatic beta cell expression of an endogenous ecoptropic proviral gene, Emy-30, located on Chr 11 of NOD and absent in the diabetes resistant NOR stock. We will establish the molecular basis for differential expression of this endogenous retrovirus in NOD beta cells versus beta cells from related but diabetes-resistant stocks. Results from these proposed studies will guide geneticists in their search for non-MHC linked diabetogenic susceptibility genes in humans and enhance understanding of their interactions with diabetogenic MHC genes.
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