Identification of Tetratricopeptide Repeat 1 as an Adaptor Protein That Interacts with Heterotrimeric G Proteins and the Small GTPase Ras

Abstract

The biological functions of heterotrimeric G proteins and small GTPases are modulated by both extracellular stimuli and intracellular regulatory proteins. Using Saccharomyces cerevisiae two-hybrid screening, we identified tetratricopeptide repeat 1 (TPR1), a 292-amino-acid protein with three TPR motifs, as a Gα16-binding protein. The interaction was confirmed both in vitro and in transfected mammalian cells, where TPR1 also binds to several other Gα proteins. TPR1 was found to interact with Ha-Ras preferentially in its active form. Overexpression of TPR1 promotes accumulation of active Ras. TPR1 was found to compete with the Ras-binding domain (RBD) of Raf-1 for binding to the active Ras, suggesting that it may also compete with Ras GTPase-activating protein, thus contributing to the accumulation of GTP-bound Ras. Expression of Gα16 strongly enhances the interaction between TPR1 and Ras. Removal of the TPR1 N-terminal 112 residues abolishes potentiation by Gα16 while maintaining the interaction with Gα16 and the ability to discriminate active Ras from wild-type Ras. We have also observed that LGN, a Gαi-interacting protein with seven TPR motifs, binds Ha-Ras. Thus, TPR1 is a novel adaptor protein for Ras and selected Gα proteins that may be involved in protein-protein interaction relating to G-protein signaling.

ACKNOWLEDGMENTS

We thank Lawrence Quilliam and Isabel Lopez for helpful discussions.

This work was made possible in part by National Institutes of Health grants AI33503 and AI40176 (to R.D.Y.). Caroline Marty is a recipient of a predoctoral fellowship from the American Heart Association Midwest Affiliate.