The general objective of the proposed research is to establish molecular mechanisms for the control of immunoglobulin (Ig) gene expression in human normal and disease states. Specifically, the organization and regulation of expression of the human lambda (Lambda) light chain V (variable), J (joining) and C (constant) gene segments will be studied. Specific objectives include the localization, isolation and characterization of JLambda gene segments, isolation of DNA probes which will detect specific VLambda subgroups and Clambda isotypes, and cloning and sequencing germline and rearranged (active) Vlambda gene segments. This project will give information on the molecular organization of the human Lambda genes, the degree and location of somatic diversification in the VLambda gene segments, which JLambda and CLambda gene segments are functional and allow molecular study of preferential V-J-C association. It will provide a basis for a molecular study of preferential light chain expression (Lambda) in particular autoimmune diseases and the preferential use of the Lambda light chain with certain heavy chain classes. An additional specific objective is to assay for transcription enhancer elements associated with the above Ig genes. For these objectives we will clone rearranged and germline Ig Lambda genes by recombinant DNA techniques, insert these genes into plasmid vectors which have positive selection markers (Ecogpt or cat), transfect various lymphoid cells with these or deleted mutants, study expression of these genes by cellular immunofluorescence and electrophoresis of proteins and nucleic acids, and sequence particular DNA fragments by established procedures.
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