Dephosphorylation of insulin-receptor autophosphorylation sites by particulate and soluble phosphotyrosyl-protein phosphatases.

Insulin stimulates autophosphorylation of the insulin receptor on multiple tyrosines in three domains: tyrosines 1316 and 1322 in the C-terminal tail, 1146, 1150 and 1151 in the tyrosine-1150 domain, and possibly 953, 960 or 972 in the juxtamembrane domain. In the present work the sequence of dephosphorylation of the various autophosphorylation sites by particulate and cytosolic preparations of phosphotyrosyl-protein phosphatase from rat liver was studied with autophosphorylated human placental insulin receptor as substrate. Both phosphatase preparations elicited a broadly similar pattern of dephosphorylation. The tyrosine-1150 domain in triphosphorylated form was found to be exquisitely sensitive to dephosphorylation, and was dephosphorylated 3-10-fold faster than the di- and monophosphorylated forms of the tyrosine-1150 domain or phosphorylation sites in other domains. The major route for dephosphorylation of the triphosphorylated tyrosine-1150 domain involved dephosphorylation of one of the phosphotyrosyl pair, 1150/1151, followed by phosphotyrosyl 1146 to generate a species monophosphorylated mainly (greater than 80%) at tyrosine 1150 or 1151. Insulin receptors monophosphorylated in the tyrosine-1150 domain disappeared slowly, and overall the other domains were completely dephosphorylated faster than the tyrosine-1150 domain. Dephosphorylation of the diphosphorylated C-terminal domain yielded insulin receptor in which the domain was singly phosphorylated at tyrosine 1322. Triphosphorylation of the insulin receptor in the tyrosine-1150 domain appears important in activating the receptor tyrosine kinase to phosphorylate other proteins. The extreme sensitivity of the triphosphorylated form of the tyrosine-1150 domain to dephosphorylation may thus be important in terminating or regulating insulin-receptor tyrosine kinase action and insulin signalling.