CELLULAR AND STRUCTURAL STUDIES OF MYELOMA MUTANTS

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My research interests focus on examining several variants of the MPC 11 IgG2b-producing mouse myeloma cell lines which synthesize altered immunoglobulin heavy chains. Some altered heavy chains are shorter than the parent, having molecular weights of 50,000 or 40,000 compared to the parental size of 55,000; others have serological, peptide and assembly characteristics of a new subclass, IgG2a. The latter are especially interesting because they represent the expression of a previously silent constant region gene. We are studying the structural relationships of these variants to each other, to the parental MPC11, and to MOPC 173, and IgG2a protein of known sequence. We have shown that the several IgG2a variant proteins retain the parental idiotype, yet differ from each other by peptide maps, charge and assembly parameters. These same parameters have enabled us to subgroup the variants. Analytical techniques of papain digestion products such as immunoelectrophoresis, SDS-PAGE of CNBr fragments, and comparative peptide maps, in addition to partial primary structural analysis have shown that the Fc of one variant protein is entirely gamma2a-like while the Fc of a second is a gammma2b-gamma 2a hybrid. Characterization of these and other variants gives us tools to localize mouse heavy chain allotypic determinants, to probe the molecular requirements for macrophage Fc receptor interactions, and to define molecular events in secretion. The findings that indicate that some variants seem to make a recombinant heavy chain and that gamma2a and gamma2b constant regions have extensive homology raise the question of whether gene rearrangements are occurring in these cells. We will prepare restriction enzyme fragments from genomic DNA of parent and variant cells and examine them by hybridization with subclass-specific cDNA probes to see if the patterns of hybridization differ. In addition, we will approach the question of the action of mutagens in these cells by asking whether after treatment with drugs such as ICR-191 or Melphalan, recombination is enhanced, or any gross chromosomal abnormalities are occurring.