Title: Tyrosine Kinase Activity of a Chimeric Insulin‐Like‐Growth‐Factor‐l Receptor Containing the Insulin Receptor C‐Terminal Domain
Abstract: In a previous study, we showed that a chimeric insulin‐like‐growth‐factor‐1 (IGF‐1) receptor, with the β subunit C‐terminal part of the insulin receptor was more efficient in stimulating glycogen synthesis and p44 mapk activity compared to the wild‐type IFG‐1 receptor [Tartare, S., Mothe, I., Kowalski‐Chauvel, A., Breittmayer, J.‐F., Ballotti, R. & Van Obberghen, E. (1994) J. Biol. Chem. 269 , 11449–11455]. These data indicate that the receptor C‐terminal domain plays an important role in the transmission of biological effects. To understand the molecular basis of the differences in receptor specificity, we studied the characteristics of insulin, IGF‐1 and chimeric receptor tyrosine kinase activities in a cell‐free system. We found that, compared to wild‐type insulin and IGF‐1 receptors, the chimeric receptor showed a decrease in (a) autophosphorylation, (b) tyrosine kinase activity towards insulin receptor substrate‐1 and the insulin re‐ceptor‐(1142–1158)‐peptide, and (c) the ability to activate phosphatidylinositol 3‐kinase. However, for all the effects measured in a cell‐free system, the chimeric receptor displayed an increased response to IGF‐1 compared to the native IGF‐1 receptor. Concerning the cation dependence of the tyrosine kinase activity, we showed that, at 10 mM Mg 2+ , the ligand‐stimulated phosphorylation of poly(Glu 80 Tyr 20 ) by both insulin receptor and chimeric receptor was increased by Mn 2+ . Conversely at 50 mM Mg 2+ , the chimeric receptor behaved like the IGF‐1 receptor, since the presence of Mn 2+ decreased the stimulatory effect of IGF‐1 on their kinase activity. Furthermore, the K m of the chimeric receptor for ATP was increased compared to the wild‐type receptors. These data demonstrate that the replacement of the C‐terminal tail of the IGF‐1 receptor by that of the insulin receptor has changed the receptor characteristics studied in a cell‐free system. Our findings indicate that the C‐terminal domain of the insulin receptor β subunit plays a key role in regulation of the tyrosine kinase activity. The fine‐tuning of the tyrosine kinase by the C‐terminal tail could participate in the receptor specificity.