An Eps Homology (EH) Domain Protein That Binds to the Ral-GTPase Target, RalBP1*
- From the Department of Biochemistry, Tufts University School of Medicine, Boston, Massachusetts 02111
Abstract
Ral proteins constitute a family of small GTPases that can be activated by Ras in cells. In the GTP-bound state, Ral proteins bind to RalBP1, a GTPase-activating protein for CDC42 and Rac GTPases. We have used the two-hybrid system in yeast to clone a cDNA for a novel ∼85-kDa protein that can bind to an additional site on RalBP1. This newly identified protein contains an Eps homology (EH) domain, which was first detected in the epidermal growth factor (EGF) receptor substrate Eps15. Recently, the EH domain of Eps15 has been shown to bind to proteins containing an asparagine-proline-phenylalanine motif. Moreover, EH domains have been found in proteins involved in endocytosis and/or actin cytoskeleton regulation. The RalBP1 associated Eps-homology domain protein, Reps1, is tyrosine-phosphorylated in response to EGF stimulation of cells. In addition, Reps1 has the capacity to form a complex with the SH3 domains of the adapter proteins Crk and Grb2, which may link Reps1 to an EGF-responsive tyrosine kinase. Thus, Reps1 may coordinate the cellular actions of activated EGF receptors and Ral-GTPases.
Footnotes
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↵* This work was supported by United States Public Health Service Grant GM47707 (to L. A. F.) from the NIGMS and an American Cancer Society senior postdoctoral grant (to T. U.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The nucleotide sequence(s) reported in this paper has been submitted to the GenBank™/EMBL Data Bank with accession number(s) AF031939.
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↵‡ These two authors contributed equally to the work.
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↵§ Present address: First Department of Surgery, Jukui Medical School, 23-4 Shimoaizuki Matsuoka-chou, Yoshida-gun, Fukui 910-11, Japan.
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↵¶ Present address: Second Department of Biochemistry, Nagoya University School of Medicine, 65 Ssurumai-chou Showaku, Nagoya 466, Japan.
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↵‖ To whom correspondence should be addressed. Tel.: 617-636-6956; Fax: 617-636-6409; E-mail: lfeig{at}opal.tufts.edu.
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↵1 The abbreviations used are: PLD, phospholipase D; GST, glutathione S-transferase; PAGE, polyacrylamide gel electrophoresis; RalBP1, Ral-binding protein 1; Reps1, RalBP1-associated Eps homology domain protein; EH, Eps homology; RACE, rapid amplification of cDNA ends; Ral-GDS, Ral-GDP dissociation stimulator; aa, amino acids.
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↵2 R. Emkey and L. A. Feig, unpublished observations.
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- Received September 17, 1997.
- Revision received October 8, 1997.
