The insulin receptor functions normally in chinese hamster ovary cells after truncation of the C terminus

Martin G. Myers, Jonathan M. Backer, Kenneth Siddle, Morris F. White

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Abstract

We studied the structure and function of the human insulin receptor (IR) and a mutant which lacked the last 43 amino acids of the β-subunit (IRΔct). This deletion removed tyrosine (Tyr1322, Tyr1316) and threonine (Thr1336) phosphorylation sites. In Chinese hamster ovary (CHO) cells, insulin binding to the mutant receptor was normal, and [35S]methionine labeling indicated that both the IR and IRΔct were processed normally; however, the β-subunit of IRΔct was 5 kDa smaller than that of the IR. The time course of insulin-stimulated autophosphorylation of the partially purified IRΔct was normal, but the maximum autophosphorylation was reduced 20-30%. Tryptic phosphopeptide mapping confirmed the absence of the C-terminal phosphorylation sites and indicated that phosphorylation of the regulatory region (Tyr1146, Tyr1150, Tyr1151) occurred normally; kinase activity of the IR and IRΔct was activated normally by insulin-stimulated autophosphorylation. In the intact CHO cells, insulin-stimulated serine and threonine phosphorylation of the IRΔct was reduced 20%, suggesting that most Ser/Thr phosphorylation sites are located outside of the C terminus. During insulin stimulation, the wild-type and mutant insulin receptor activated the phosphatidylinositol 3-kinase. Moreover, insulin itself or human-specific antiinsulin receptor antibodies stimulated glycogen and DNA synthesis equally in both CHO/IR and CHO/IRΔct cells. These data suggest that the C terminus plays a minimal role in IR function and signal transmission in CHO cells.

Original languageEnglish (US)
Pages (from-to)10616-10623
Number of pages8
JournalJournal of Biological Chemistry
Volume266
Issue number16
StatePublished - 1991
Externally publishedYes

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Insulin Receptor
Cricetulus
Phosphorylation
Ovary
Cells
Insulin
Threonine
Phosphatidylinositol 3-Kinase
Phosphopeptides
Nucleic Acid Regulatory Sequences
Glycogen
Methionine
Labeling
Serine
Tyrosine
Phosphotransferases
Amino Acids
Antibodies
DNA

ASJC Scopus subject areas

  • Biochemistry

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The insulin receptor functions normally in chinese hamster ovary cells after truncation of the C terminus. / Myers, Martin G.; Backer, Jonathan M.; Siddle, Kenneth; White, Morris F.

In: Journal of Biological Chemistry, Vol. 266, No. 16, 1991, p. 10616-10623.

Research output: Contribution to journalArticle

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