The proposed experiments will address the problem of the ontogeny of T lymphocytes in adult and developing mice. Several new techniques based on retroviral-mediated gene transfer will be combined with the established methods of monoclonal antibody cell surface markers to examine the general questions of lineage relationships within the T cell compartment, tissue-specific expression of genes by T cell progenitors and immature T cells, and dynamics of T cell maturation during development. Replication-defective retroviruses will be used to infect murine bone marrow cells and these cells will be introduced into irradiated recipients. Each infection event by virtue of its unique position of chromosome insertion marks a stem cell and its progeny. Analysis of animals at various times after transplantation will allow estimation of numbers of progenitors involved in thymic and peripheral T cell repopulation. The distribution of clonal markers in monoclonal antibody-defined thymocyte subpopulations will also be studied. These experiments may allow the dissection of separate intrathymic lineages and inferences on ordered precursor-progeny relationships. Second, novel retrovirus vectors will be constructed with the goal of high-efficiency expression of transferred genes in thymocytes. Combination of this method with introduction of a transforming gene, SV40 large T-antigen, under the control of the adenosine deaminase (ADA) promoter may allow clonal expansion of the thymus subpopulation having high ADA expression. Third, by introduction of retroviruses into developing mouse fetuses, lymphocyte stem cell migration and cell fates will be explored. Use of a novel marker gene, E. coli lacZ, should allow in situ examination of marked thymocyte stem cells. Control of gene expression after retrovirus infection of postimplantation embryos will also be explored. These studies should allow definition of lineage relationships and stem cell regulation in the T cell compartment not heretofore possible. Better understanding of T cell development should have wide ranging implications for studies of immune function in genetic immune deficiencies, response to infectious agents, and in autoimmune disorders.
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