Insulin receptor substrate-1 mediates phosphatidylinositol 3'-kinase and p70S6k signaling during insulin, insulin-like growth factor-1, and interleukin-4 stimulation.

M.G Myers,T.C Grammer,L.M Wang,X.J Sun,J.H Pierce,J Blenis,M.F White

doi:10.1016/s0021-9258(19)61974-5

Abstract

Insulin Receptor Substrate-1 (IRS-1) is an endogenous cellular protein that is tyrosine phosphorylated during stimulation of cells with insulin, IGF-1, and interleukin 4 (IL-4). Phosphorylated IRS-1 regulates multiple regulatory pathways by recruiting signaling molecules containing Src homology 2 domains (SH2 proteins). The 32D myeloid progenitor cell line contains few insulin receptors and no detectable IRS-1. Expression of the insulin receptor alone partially mediates insulin-stimulated microtubule-associated protein (MAP) kinase activation, and the addition of IRS-1 enhances this effect (Myers, M. G., Jr., Wang, L.-M., Sun, X. J., Zhang, Y., Yenush, L. P., Schlessinger, J., Pierce, J. H., and White, M. F. (1994) Mol. Cell. Biol. 14, 3577-3587). Alone, insulin receptors mediate phosphatidylinositol (PI) 3'-kinase and p70S6k activation poorly if at all during insulin stimulation. Expression of IRS-1 alone in 32D cells mediates the stimulation of p70S6k by insulin, IGF-1, or IL-4; addition of insulin receptor to these cells increases the sensitivity of the insulin response. In contrast, full insulin stimulation of PI 3'-kinase requires both the insulin receptor and IRS-1, suggesting that a high level of IRS-1 phosphorylation is required for insulin-stimulated PI 3'-kinase activation, whereas a low level of IRS-1 tyrosine phosphorylation transmits an essential signal to p70S6k. Both insulin receptors and IRS-1 are required for mitogenic signaling in 32D cells suggesting that MAP kinase or p70S6k alone are not sufficient, and that both or additional unknown IRS-1-mediated signals are necessary.

Highlights

Insulin Receptor Substrate-1(IRS-1) iasn endogenous (SH2) domains [1,2,3]
IRS-1 interacts directly with the insulin cellular protein that is tyrosine phosphorylated during receptor and undergoes tyrosine phosphorylation during insustimulation of cells withinsulin, IGF-1, and interleukin lin stimulation[1,2,3]
The32D myeloid progenitor cell line contains low levels of insulin receptor and no detectable IRS-1 or related molecules [11].the signalsmediated by the insulin receptor can be investigated in the presence and sion of IRS-1 alone i3n2D cells mediatesthe stimulation absence of IRS-1 in 32D cells, and the signaling pathwaycsonof ~ 7 0 % by ~insulin, IGF-1, or IL-4; addition of insulin trolled by IRS-1 can be clearly defined

Summary

88 To whom correspondence and reprint requests should be addressed

Research Division, J o s h DiabetesCenter, 1 JoslinPlace, Boston, MA 02215.Tel.:617-732-2578;Fax:617-732-2593;E-mail: the insulin receptor [1]. The insulinreceptor directly binds andactivates PI 3”kinase in vitro [23], much of the receptor-associated PI 3”kinase activity i n vivo represents the association of the PI 3’-kinase/IRS-1 complex with the acti-. Like other growth factors, activates a network of serinelthreonine kinases, including the MAP kinase/p9Wkcascade, and ~ 7 [260-28]. The insulin receptor appears to engage the upstream elements of the MAP/p9Orskcascade (including p21H-"") through the phosphorylation of IRS-1 or Shc which associate with Grb-S/SOS[7,29,30]. Thymidine incorporation into DNA in 32D cell lines. Data are the average of triplicate determinations graphed as thepercent of the positive control (IL-3-stimulated thymidine incorporation).

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION