Mousepox is an important viral disease of laboratory mice with remarkably varied symptomatology based on the presence or absence of multiple resistance genes. The long term objectives of this research are to identify, localize and clone the major mousepox resistance genes, to determine their mechanisms of action, and to devise strategies based on transgenic technology to protect susceptible strains from ectromelia virus. For the major mousepox resistance genes, this application proposes to complete development of congenic strains, to continue defining mechanisms of action, to complete chromosomal localization, and to begin fine mapping. Resistance is determined prior to the onset of specific immunity, requires natural killer (NK) cells, and one resistance gene, RMP-1, maps near a complex of genes, NKR-P1, including that which encodes NK 1.1, expressed solely by NK cells. To investigate the role of NK cells in resistance, mixed chimeras will be produced which carry NK cells from resistant NK 1.1+ (Rmp-1r) and susceptible NK 1.1- congenic donors. Three methods will be used to produce mixed chimeras, parabiosis, adoptive transfer of adult spleen cells to infants, and radiation bone marrow chimeras. The method that best demonstrates transfer of resistance to the susceptible congenic partner will be used. The effect of ablation of NK 1.1+ cells in NK 1.1+/NK1.1- chimeras by anti-NK 1.1 antibody on virus replication will determine if Rmp-1 is expressed by NK 1.1+ cells. If Rmp-1 is expressed by NK 1.1+ cells, a modification of this protocol will be used to determine if other resistance genes are expressed by NK cells. The resistant donors will be F1 hybrids of the Rmp-1r congenic strain and congenic strains for the other resistance genes of interest. Because F1 cells will express both dominant resistance genes and NK 1.1, ablation of NK 1.1+ cells will augment virus replication if the other resistance genes are also expressed by NK cells. To determine if resistance genes that are not expressed by NK cells act through NK cells, levels of NK cell expression of NKR-P1 genes and immune interferon (IFN-gamma), which exhibits sex-biased expression like some resistance genes, will be tested in ectromelia infected congenic strains.The complement system and T cell precursors have also been implicated in resistance to mousepox.To determine if those congenic resistant strains with genes that are not expressed by or through NK cells mediate resistance through complement of T cell precursors. The effects of complement and T cell depletion on virus replication will be examined. The chromosomal location of Rmp-4, the only unlocalized resistance gene, will be determined by multilocus analysis of the Rmp-4 congenic strain and the susceptible congenic partner using probes for mapped non ecotropic proviruses. The four resistance genes will be fine mapped by determining loci on the donor segments of congenic resistant strains and using these loci in linkage analysis with resistance genes in backcross mice.
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