HUMAN INSULIN-RECEPTOR MUTATED AT THREONINE 1336 FUNCTIONS NORMALLY IN CHINESE-HAMSTER OVARY CELLS

Phosphorylation of threonine 1336 of the human insulin receptor (HIR) is stimulated by insulin or 4beta-phorbol 12-myristate 13-acetate in C hinese hamster ovary (CHO) transfectant cells expressing the wild type receptor (CHO/HIR). To examine the role of this phosphorylation in in sulin signal transduction, a mutant human insulin receptor, in which t hreonine 1336 was replaced with asparagine, has been stably expressed in CHO cells (CHO/HIRT1336N). CHO cell lines expressing equivalent num bers of the wild type or the mutant receptor were developed, which bou nd I-125-insulin comparably (K(d) = 0.1 nM). After stimulation of CHO/ HIR or CHO/HIRT1336N cells with insulin, the wild type and mutant rece ptors internalized the hormone and were down-regulated with similar ra tes. Hormone stimulation of the receptor tyrosine kinase activity was also unaffected by the mutation. Metabolic and mitotic effects of insu lin were also unimpaired by the mutation. Thus, insulin stimulated pho sphatidylinositol 3-kinase activity, glycogen synthesis, and thymidine incorporation into DNA similarly in CHO/HIR and CHO/HIRT1336N cells. These data suggest that by itself phosphorylation of threonine 1336 ha s no significant effect on insulin binding, regulation of insulin rece ptor expression, or insulin signal transduction.