Deletion of C-terminal 113 amino acids impairs processing and internalization of human insulin receptor: comparison of receptors expressed in CHO and NIH-3T3 cells.
We have studied the structure and the function of a truncated human insulin receptor in which 113 amino acids (aa 1231-1343) at the C-terminus of the beta-subunit were deleted. In this study, wild-type and truncated insulin receptors were expressed by stable transfection in NIH-3T3 cells and CHO cells. The mutation impairs post-translational processing of the insulin receptor; proteolytic cleavage is retarded, and degradation of the truncated receptor is accelerated. Furthermore, insulin-stimulated autophosphorylation of the mutant insulin receptor is impaired. This is associated with a defect in insulin-stimulated endocytosis. Finally, in NIH-3T3 cells, the mutant insulin receptor failed to mediate the mitogenic effects of insulin. In CHO cells, transfection of insulin receptor cDNA (either wild-type or mutant) did not alter mitogenic response to insulin. It has previously been shown that deletion of 43 amino acids at the C-terminus of the beta-subunit did not affect insulin receptor tyrosine kinase activity. Our data suggest that the structural domain located 43-113 amino acids from the C-terminus appears to have several functional roles. First, the domain appears to promote folding of receptor into the optimal conformation for post-translational processing. Second, the presence of this domain appears to promote the stability of the receptor beta-subunit in intact cells. Finally, perhaps as a consequence of the effects upon the stability of the receptor, this domain is required in intact cells for insulin-stimulated autophosphorylation and signal transmission.
Levy-Toledano
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Diabetes Branch, National Institute of Diabetes and Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892.
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Taylor
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D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D004247:DNA; D004705:Endocytosis; D015870:Gene Expression; D006801:Humans; D007328:Insulin / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D011972:Receptor, Insulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D013936:Thymidine / Q000378:metabolism; D014162:Transfection
D007328:Insulin; D010446:Peptide Fragments; D004247:DNA; D011972:Receptor, Insulin; D013936:Thymidine