Infection of bone marrow on fetal liver with Abelson Murine Leukemia virus (A-MuLV) transforms a small fraction of the cells into clonal continuous cell lines which derive from B-lymphoid (immunoglobulin-producing) cells. We have defined several sub-classes of these transformants which in culture undergo the molecular rearrangements associated with immunodifferentiation. The work described in this proposal is aimed at exploiting the properties of these sub-classes to study the regulation of Ig gene rearrangement and expression. Most previous studies in this area have focused on cell lines which represented the later stages of the B-cell pathway (i.e., myelomas) and which were terminally reorganized with respect to Ig gene structure. In contrast, most of our proposed studies will focus on cell lines which, in culture, undergo the rearrangement processes associated with the construction of complete heavy and light chain genes. With such lines we propose to analyze, in more detail than previously possible, the mechanism by which a given B-cell clone achieves functional rearrangement and expression of only one of its two heavy chain alleles (allelic exclusion) and one of its multiple light chain genes (allelic and isotypic exclusion). Details of the rearrangement process will be analyzed by the introduction of appropriate recombination substrates into cells which are actively undergoing heavy or light chain gene rearrangement. The hypothesis that these rearrangement processes are controlled by feedback from Ig chains will be tested by introduction of functional Ig genes into these lines -- both by cell infusion and gene transfer protocols. Finally, additional studies will focus on the mechanism by which expression of the functionally rearranged heavy chain gene is regulated over the course of B-cell differentiation. The availability of variant lines of A-MuLV tranformants with altered regulation of heavy chain gene expression should greatly facilitate these studies.
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