Cross-linking of insulin receptors to MHC antigens in human B lymphocytes: evidence for selective molecular interactions.
Molecular interactions between insulin receptors and MHC antigens were investigated in human B cells. Two B lymphoblastoid cell lines, IM-9 and 526, chosen for their high insulin binding capacity, were found to express 15,000 and 25,000 insulin receptors per cell, respectively. Insulin receptors were labeled with a 125I-photoreactive insulin analogue, and all other surface proteins by lactoperoxidase-catalyzed radioiodination. Neighbor proteins were cross-linked with a cleavable homobifunctional reagent dithio-bis-(succinimidyl propionate) (DSP) and solubilized before immunoprecipitation by anti-HLA monoclonal antibodies. Gel analysis of the precipitated proteins showed that 90% of insulin receptors precipitable by anti-insulin receptor antibodies were precipitated by anti-class I antibodies (anti-heavy chain and anti-beta 2-microglobulin) after cross-linking with 2 mM DSP. In neither IM-9- nor 526 cells could HLA antigens be precipitated by anti-insulin receptor antibodies, suggesting that the concentration of class I antigens largely exceeds the concentration of insulin receptors at the cell surface. In 526 lymphocytes, class I MHC antigens were also found to adjoin class II antigens, since both molecules could be coprecipitated with anti-HLA A, B, C and with anti-HLA-DR antibodies after chemical cross-linking. Down-regulation of insulin receptors by chronic exposure of IM-9 cells to insulin did not affect the amount of MHC molecules present on the cell surface, and conversely, class I MHC molecules were internalized in 526 cells irrespective of the presence of insulin. These results thus show that insulin receptors and MHC antigens form multimolecular complexes in the plasma membrane of cultured human B cells. These interactions, which do not appear to influence the regulation of these proteins on the cell surface, may be involved in the mechanism of hormone signaling.
D001402:B-Lymphocytes / Q000648:ultrastructure*; D003432:Cross-Linking Reagents; D004705:Endocytosis; D006680:HLA Antigens; D006681:HLA-D Antigens; D006801:Humans; D007152:Immunologic Capping; D008285:Major Histocompatibility Complex; D011972:Receptor, Insulin / Q000378:metabolism*; D001613:beta 2-Microglobulin / Q000378:metabolism
D003432:Cross-Linking Reagents; D006680:HLA Antigens; D006681:HLA-D Antigens; D001613:beta 2-Microglobulin; D011972:Receptor, Insulin