Abstract
We constructed mutant receptors by mutating transmembrane Val922 of the human insulin-like growth factor I receptor (IGF-IR). Assays of receptor kinase and autophosphorylation revealed constitutively augmented tyrosine kinase activity of V922E IGF-IR in both transient and stable expression. The constitutively active tyrosine kinase of this mutant was verified by promoted tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) in the absence of IGF-I. In CHO cells stably increasing V922E IGF-IR, both IRS-1 phosphorylation and the IRS-1 associated phosphoinositide 3-kinase activity were stimulated in the absence of IGF-I to the level attained by 1 nM IGF-I stimulation of wild-type IGF-IR, whereas the Ras-mitogen-activated protein kinase pathway was not activated under the same condition. In these CHO cells, V922E IGF-IR significantly stimulated glucose uptake but did not promote mitogenesis in the absence of IGF-I. We thus conclude that the V922E mutation of IGF-IR switches on the intrinsic tyrosine kinase and differentially activates the downstream pathways. This mutant is extremely useful in clarifying the turning-on mechanism of IGF-IR as well as the differential roles of individual downstream pathways of receptor tyrosine kinases.
Highlights
From the Cardiovascular Research Center, Massachusetts General Hospital-East and the Department of Medicine, Harvard Medical School, Charlestown, Massachusetts 02129
In CHO cells stably expressing V922E IGF-IR, both illS·1 phosphorylation and the insulin receptor substrate-I (IRS-I)-associated phosphoinositide 3-kinase activity were stimulated in the absence of IGF-I to the level attained by I nM IGF-I stimulation of wild-type IGF-IR, whereas the Ras-mitogen-activated protein kinase pathway was not activated under the same condition
IGF-IRs transiently expressed in COS-7 cells treated with or without 100 nM IGF-I were precipitated with aIR3
Summary
Ways by recruiting PI 3-kinase [8, 9], tyrosine-specific protein phosphatase Syp [10], and probably others. The downstream pathways ofIGF-IR so far clarified are all shared by IR [11] It remains unknown if each receptor has its unique signaling pathway or if such differences result from the different activation profiles of the same downstream pathways. To address these questions, it is necessary to turn on IGF-IR selectively. One of the best ways is to construct and use a constitutively active mutant of IGF-IR. The expression of such an autoactive mutant allows the precise analysis of the downstream pathways and functional roles of IGF-IR. Differential roles of IGF-IR signaling pathways in biological outputs are discussed by comparing the metabolic and mitogenic effects of this activated mutant to its intracellular signals
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
