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 4 beta-phorbol 12-myristate 13-acetate in Chinese hamster ovary (CHO) transfectant cells expressing the wild type receptor (CHO/HIR). To examine the role of this phosphorylation in insulin signal transduction, a mutant human insulin receptor, in which threonine 1336 was replaced with asparagine, has been stably expressed in CHO cells (CHO/HIRT1336N). CHO cell lines expressing equivalent numbers of the wild type or the mutant receptor were developed, which bound 125I-insulin comparably (Kd = 0.1 nM). After stimulation of CHO/HIR or CHO/HIRT1336N cells with insulin, the wild type and mutant receptors internalized the hormone and were down-regulated with similar rates. Hormone stimulation of the receptor tyrosine kinase activity was also unaffected by the mutation. Metabolic and mitotic effects of insulin were also unimpaired by the mutation. Thus, insulin stimulated phosphatidylinositol 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 has no significant effect on insulin binding, regulation of insulin receptor expression, or insulin signal transduction.
Ahn
J
J
Memorial Sloan-Kettering Cancer Center, New York, New York.
Rosen
O M
OM
Donner
D B
DB
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D004247:DNA; D015536:Down-Regulation; D004789:Enzyme Activation; D005947:Glucose / Q000378:metabolism; D006003:Glycogen / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism; D011972:Receptor, Insulin / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D013912:Threonine / Q000378:metabolism*
D007328:Insulin; D013912:Threonine; D006003:Glycogen; D004247:DNA; D010770:Phosphotransferases; D019869:Phosphatidylinositol 3-Kinases; D011972:Receptor, Insulin; D005947:Glucose