The long-term objective of this work are two-fold: 1. To ascertain the role, in pre-B lymphocytes, of intracellular membrane immunoglobulin heavy chain protein (mum) and two pre-B specific surrogate Ig light chain proteins w and L, with particular reference to the feedback regulation of B cell differentiation. The clonal specificity of B lymphocytes is maintained by feedback processes which ensure allelic and isotopic exclusion. Although dire immunological consequences may be predicted if individual lymphoid clones expressed multiple receptors, human disorders resulting from aberrant feedback regulation processes have as yet to be identified. 2. To obtain a molecular understanding of the cellular pre-B to B cell transition with particular reference to the B cell specific transport of mum to the cell surface and the shut-off of surrogate light chain gene expression. A detailed understanding of events at this transition will be of direct relevance to X-linked agammaglobulinemia, a human immunodeficiency disease in which cells remain frozen at the pre-B stage. The surrogate light chain proteins, omega and iota, as well as 6 polypeptides associated in pre-B cell with mum2omega2 and mum2iota2 tetrameres, will be purified and sequenced. Antibodies against these proteins will be generated. The genes that encode these proteins will be cloned and expressed in tissue culture lines for studies relevant to both the structure of these proteins and their function during differentiation. Site directed mutagenesis of the mum gene will be employed to determine the function of myristylation in the B stage specific transport of mum to the cell surface. A search for dextransulfate induced genes that regulate mum transport will be conducted using a transfection assay system in a fibroblast cell line. The role of """"""""conventional"""""""" light chain protein in signalling the shut-off of surrogate light-chain gene expression will be explored.
