The recognition of antigen by the MHC-restricted antigen receptor on T lymphocytes can result in a series of interrelated events culminating in entry into the cell cycle, DNA synthesis, and clonal expansion by mitotic division. The magnitude of T cell activation following exposure to T cell mitogens segregates as a genetic trait in rodents. This trait is most robust in inbred rats, where there can be a 100-200 fold difference in thymidine uptake between high (Lewis) and low (BN) responder strains. Low responders can be converted to high responders in vitro through the addition of nitric oxide synthase inhibitors. Previous backcross analyses between low and high responder strains suggested that this trait segregated as an autosomal trait with high response to mitogen dominant, and an inheritance pattern consistent with a single gene. However, this latter conclusion was based on small numbers of animals. Therefore, in our first Specific Aim, we propose to define the mode of inheritance of this trait. We will breed and phenotype the offspring of the following crosses: (Lewis x BN)F1 x BN and (Lewis x BN)F2. Phenotypic analysis consists of activating their splenocytes with T cell mitogens in the presence and absence of a selective iNOS inhibitor. In the second Specific Aim, we propose to utilize SSLP markers from the rat genome map to define quantitative trait loci important for controlling the ability to proliferate to mitogens. We will use genomic DNA isolated from livers of the animals that have been phenotyped for their proliferative responses. Pools of DNA from high and low responder animals will be scanned utilizing polymorphic markers placed at 10-15 cM intervals that distinguish Lewis and BN rats. Suggestive pools will be meticulously analyzed by examining individual animals. The Mapmaker/QTL program will be used to identify SSLPs related to this phenotypic trait. We will localize the trait to an interval of approximately 1 centimorgan. In the final specific aim, we will return to two experimental autoimmune systems, experimental allergic encephalomyelitis and anti-tubular basement membrane disease with interstitial nephritis, to examine whether the locus responsible for this proliferative response alters the severity of autoimmune disease. We propose to make and utilize two different """"""""speed"""""""" congenics, one in which the BN QTL is expressed on the Lewis background, and one in which the Lewis QTL is expressed on the BN background for these studies. These studies will provide important new information regarding genes that regulate the magnitude of lymphocyte responses and susceptibility/resistance to autoimmune disease.
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