Biomedical subjects
Publications and source records attributed to I Mellman.
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We have studied the intracellular binding of radioactive cobalamin by normal cultured human fibroblasts grown in medium containing [(57)Co]-cobalamin. We have also assessed the significance of defects in this binding activity exhibited by two classes of human mutants (cbl C and cbl D) each characterized by pleiotropic deficiencies in the accumulation and retention of cobalamin, in the synthesis of cobalamin coenzymes, and accordingly, in the holoenzyme activities of both cobalamin-dependent enzymes, 5-methyltetrahydrofolate:homocysteine methyltransferase and methylmalonyl-CoA mutase. Based on the coincidence of [(57)Co]cobalamin binding and cobalamin-dependent enzyme activities after Sephadex G-150 chromatography and polyacrylamide gel electrophoresis, we conclude that, as in rat liver, the intracellular binding of labeled cobalamin by normal fibroblasts reflects the attachment of the vitamin to the cobalamin-dependent methyltransferase and mutase. Whereas cbl C cells are completely deficient in the binding of [(57)Co]cobalamin to either enzyme, fibroblasts which bear the phenotypically similar but genetically distinct cbl D mutation retain some binding activity, and accordingly, have higher holomethyltransferase and holomutase activities than do cbl C cells. The defect in [(57)Co]-cobalamin binding exhibited by both cbl C and cbl D fibroblasts is almost certainly not a result of mutations which affect the methyltransferase or mutase apoenzymes, since the electrophoretic mobilities and the affinities of these enzymes for their respective cobalamin coenzymes are indistinguishable from those in control cell extracts. These results suggest that both the cbl C and cbl D mutations affect some enzymatic step(s) which converts newly taken up cobalamin to a form capable of being bound by the two cobalamin-dependent enzymes.
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Macrophages, granulocytes and many lymphocytes express or secrete receptors for the Fc domain of immunoglobulins (Ig). These Fc receptors (FcRs) are heterogeneous and can be distinguished on the basis of their cellular distribution and specificities for different immunoglobulin isotypes. Although their functions are not completely understood, FcRs are known to be involved in triggering various effector cell functions and in regulating differentiation and development of B-cells. One of the best characterized is the mouse macrophage-lymphocyte receptor for IgG1 and IgG2b (ref. 5). On macrophages, this FcR mediates the endocytosis of antibody-antigen complexes via coated pits and coated vesicles, the phagocytosis of Ig-coated particles, and the release of various inflammatory and cytotoxic agents. It is possible that the receptor possesses an intrinsic ligand-activated ion channel activity responsible for some of these functions. The IgG1/IgG2b FcR has been isolated and shown to be a transmembrane glycoprotein of relative molecular mass (Mr) 47,000-60,000 (47-60 K) containing four N-linked oligosaccharide chains and a large (greater than 10K) cytoplasmic domain. It is also immunologically indistinguishable from the murine Ly-17 alloantigen which, in turn, is tightly linked to the Mls lymphocyte activation locus. Here we describe the isolation and characterisation of a complementary DNA clone encoding the whole of the IgG1/IgG2b FcR expressed by the mouse macrophage-like cell line P388D1. The receptor is a member of the immunoglobulin superfamily and, like Ly-17, maps to the distal portion of chromosome 1. cDNA probes detect one or two mRNA species in FcR+ macrophage and B-cell lines, but not in FcR- cells or a receptor-deficient variant derived from a FcR+ B-cell line. Finally, DNA hybridization analysis indicates the receptor gene is partially deleted or rearranged in the FcR- variant.