The p120 GTPase-activating protein (GAP) is a negative regulator of Ras, which has a central role in signal transduction pathways that control cell proliferation. p120 GAP accelerates the conversion of activated Ras-GTP to its inactive form, Ras-GDP, thereby inhibiting mitogenic signaling. To examine potential contributions of p120 N-terminal sequences to regulation of its C-terminal catalytic domain, we constructed deletion mutants lacking defined regions, including the variable hydrophobic region as well as the Src homology 2 (SH2) and 3 (SH3) domains. These mutant proteins were expressed in infected Sf9 insect cells from recombinant baculoviruses and assayed in vitro for their ability to stimulate the intrinsic GTPase activity of purified Ras. While deletion of the variable hydrophobic region had no effect on p120 GAP activity, deletion of the entire SH2/SH3/SH2 region severely impaired catalytic activity toward Ras. Deletion of individual SH2 and SH3 domains within this region partially inhibited p120 GAP activity. Moreover, p120 N-terminal sequences enhanced the Ras GTPase-stimulating activity of the neurofibromin GAP-related domain. These results demonstrate that sequences in the SH2/SH3/SH2 region of p120 GAP are required for full catalytic activity toward Ras. Together with earlier findings that the p120 GAP SH2 domains mediate interactions with several GAP-associated proteins, our results suggest multiple roles for the N-terminal sequences in regulating p120 GAP catalytic activity and mitogenic signaling pathways. In addition, our results raise the possibility that SH2 domain point mutations in p120 GAP detected in some basal cell carcinomas reduce catalytic activity toward Ras and thereby contribute to oncogenesis.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

This article has been cited by other articles:

				P. Lock, F. Casagranda, and A. R. Dunn Independent SH2-binding Sites Mediate Interaction of Dok-related Protein with RasGTPase-activating Protein and Nck J. Biol. Chem., August 6, 1999; 274(32): 22775 - 22784. [Abstract] [Full Text] [PDF]

				I.-e. Gallouzi, F. Parker, K. Chebli, F. Maurier, E. Labourier, I. Barlat, J.-P. Capony, B. Tocque, and J. Tazi A Novel Phosphorylation-Dependent RNase Activity of GAP-SH3 Binding Protein: a Potential Link between Signal Transduction and RNA Stability Mol. Cell. Biol., July 1, 1998; 18(7): 3956 - 3965. [Abstract] [Full Text]

				B. Zhang, Z.-X. Wang, and Y. Zheng Characterization of the Interactions between the Small GTPase Cdc42 and Its GTPase-activating Proteins and Putative Effectors. COMPARISON OF KINETIC PROPERTIES OF Cdc42 BINDING TO THE Cdc42-INTERACTIVE DOMAINS J. Biol. Chem., August 29, 1997; 272(35): 21999 - 22007. [Abstract] [Full Text] [PDF]

				S. Li, S. Nakamura, and S. Hattori Activation of R-Ras GTPase by GTPase-activating Proteins for Ras, Gap1m, and p120GAP J. Biol. Chem., August 1, 1997; 272(31): 19328 - 19332. [Abstract] [Full Text] [PDF]