Direct Association of Grb2 with the p85 Subunit of Phosphatidylinositol 3-Kinase

doi:10.1074/jbc.270.21.12774

Direct Association of Grb2 with the p85 Subunit of Phosphatidylinositol 3-Kinase(*)

From the (1) Division of Cell and Molecular Biology, Dana-Farber Cancer Institute and the
(2) Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115

^§ To whom correspondence should be addressed:
Dept. of Pathology, Harvard Medical School, Dana-Farber Cancer Inst., M 857, 44 Binney Street, Boston, MA 02115.
Tel.: 617-632-3049; Fax: 617-632-4770.

Abstract

Phosphatidylinositol 3-kinase (PI 3-kinase) has been shown to play a key role in growth factor signaling pathways, although its signaling mechanism has not been fully elucidated. Using the yeast interaction trap system, we have identified Grb2 as a PI 3-kinase interacting protein. Our experiments demonstrate that p85, the regulatory subunit of PI 3-kinase, interacts with Grb2 in vivo, and this interaction is independent of growth factor stimulation. The direct association between Grb2 and p85 was reconstituted in vitro with glutathione S-transferase fusion proteins. Domain analyses and peptide competition indicate that the association is mediated by the SH3 domains of Grb2 and the proline-rich motifs of p85 and that only one SH3 domain is required for minimal binding. The interaction does not displace the catalytic subunit of PI 3-kinase but is exclusive of Sos. Signaling through PI 3-kinase, therefore, may involve the ubiquitous adapter Grb2, which serves as a convergence point for multiple pathways.

Footnotes

↵* This work was supported by National Institutes of Health Grant CA30002 and a Sandoz Fellowship (to J. W.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

PI 3-kinase

phosphatidylinositol 3-kinase

bcr

break point region

GST

glutathione S-transferase

PDGF

platelet-derived growth factor

PBS

phosphate-buffered saline

PAGE

polyacrylamide gel electrophoresis

SH

Src homology domain

x-gal

5-bromo-4-chloro-3-indoyl β-D-galactoside.
↵²K. R. Auger, L. Jarvis, and T. M. Roberts, unpublished results.