The carboxyl terminal extension of the Drosophila insulin receptor homologue binds IRS-1 and influences cell survival.
The Drosophila insulin receptor (INR) homolog includes an extension of approximately 400 amino acids at the carboxyl-terminal end of its beta subunit containing several tyrosine-based motifs known to mediate interactions with signaling proteins. In order to explore the role of this extension in INR function, mammalian expression vectors encoding either the complete INR beta subunit (beta-Myc) or the INR beta subunit without the carboxyl-terminal extension (betaDelta) were constructed, and the membrane-bound beta subunits were expressed in 293 and Madin-Darby canine kidney cells in the absence of the ligand-binding alpha subunits. beta-Myc and betaDelta proteins were constitutively active tyrosine kinases of 180 and 102 kDa, respectively. INR beta-Myc co-immunoprecipitated a phosphoprotein of 170 kDa identified as insulin receptor substrate-1 (IRS-1), whereas INR betaDelta did not, suggesting that the site of interaction was within the carboxyl-terminal extension. IRS-1 was phosphorylated on tyrosine to a much greater extent in cells expressing INR beta-Myc than in parental or INR betaDelta cells. Despite this, a variety of PTB or SH2 domain-containing signaling proteins, including IRS-2, mSos-1, Shc, p85 subunit of phosphatidylinositol 3-kinase, SHP-2, Raf-1, and JAK2, were not associated with the INR beta-Myc.IRS-1 complex. Overexpression of INR beta-Myc and betaDelta kinases conferred an equivalent increase in cell proliferation in both 293 and Madin-Darby canine kidney cells, indicating that this growth response is independent of the carboxyl-terminal extension. However, INR beta-Myc-expressing cells exhibited enhanced survival relative to parental and betaDelta cells, suggesting that the carboxyl-terminal extension, through its interaction with IRS-1, plays a role in the regulation of cell death.
Marin-Hincapie
M
M
Department of Anatomy and Cell Biology, State University of New York, Health Science Center, Brooklyn, New York 11203, USA.
Garofalo
R S
RS
D000818:Animals; D002455:Cell Division; D002460:Cell Line; D002470:Cell Survival; D004285:Dogs; D004330:Drosophila / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D055504:Insulin Receptor Substrate Proteins; D009154:Mutation; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D011972:Receptor, Insulin / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection; D018909:src Homology Domains
C526292:IRS1 protein, human; D055504:Insulin Receptor Substrate Proteins; D010750:Phosphoproteins; D019000:Phosphotyrosine; D011972:Receptor, Insulin