Functional Interaction between c-Src and Its Mitotic Target, Sam 68

doi:10.1074/jbc.270.17.10120

Functional Interaction between c-Src and Its Mitotic Target, Sam 68 (*)

From the (1) Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853 and the
(2) Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario M5G 1X5, Canada

^§ To whom correspondence and reprint requests should be addressed. Tel.: 607-254-4896; Fax: 607-255-2428.

Abstract

The c-Src tyrosine kinase phosphorylates and binds to a 68-kDa RNA-binding protein in mitotic cells. We have examined the mechanism and functional consequence of the interaction of c-Src with this protein, Sam 68 (Src associated in mitosis, 68 kDa). In whole cell homogenates, Sam 68 was the predominant substrate and binding partner of overexpressed c-Src. Mitotic, tyrosine-phosphorylated Sam 68 bound selectively to recombinant SH2 domains with significantly different affinities (c-Src ≈ Ras GTPase activating protein > p85α (amino-terminal) > Grb2 p85α (COOH-terminal)). In vitro translated Sam 68 also bound selectively to recombinant SH3 domains, with the highest affinity for the Src and p85α SH3 domains. SH3 binding was inhibited by specific Sam 68 peptides. In vitro translated Sam 68 bound directly to immobilized poly(U), and this was inhibited by binding of Src and p85 SH3 domains to Sam 68. The results suggest that the selection of Sam 68 as a mitotic target by c-Src is the result of highly specific interaction with SH2 and SH3 domains and that this interaction may modulate the RNA binding activity of Sam 68.

Footnotes

↵* This research was supported by National Institutes of Health Grant CA 32317 (to D. S.). 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:

Sam 68

Src associated in mitosis (68 kDa)

GAP

GTPase-activating protein

GST

glutathione S-transferase

PTyr

phosphotyrosine.
↵²S. J. Taylor and D. Shalloway, unpublished observations.
↵³S. J. Taylor and D. Shalloway, unpublished results.