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RAFTK/Pyk2 tyrosine kinase mediates the association of p190 RhoGAP with RasGAP and is involved in breast cancer cell invasion

Oncogene volume 19, pages 1318–1328 (2000)Cite this article

Abstract

Focal adhesions and actin cytoskeleton are involved in cell growth, shape and movement and in tumor invasion. Mitogen-induced changes in actin cytoskeleton are accompanied by changes in the tyrosine phosphorylation of several focal adhesion proteins. In this study, we have investigated the role of RAFTK, a cytoplasmic tyrosine kinase related to focal adhesion kinase (FAK), in heregulin-mediated signal transduction in breast cancer cells. Stimulation of T47D cells with heregulin (HRG) induced the tyrosine phosphorylation of RAFTK and the formation of a multiprotein complex. Analyses of the members of the HRG-stimulated complex revealed that RAFTK is associated with p190 RhoGAP (p190), RasGAP and ErbB-2, and plays an essential role in mediating the tyrosine phosphorylation of p190 by Src. Mutation of the Src binding site within RAFTK (402) abolished the phosphorylation of p190. In addition, upon HRG stimulation of T47D cells, association of ErbB-2 with RAFTK was observed and found to be indirect and mediated by Src. Expression of wild-type RAFTK (WT) significantly increased MDA-MB-435 and MCF-7 breast cancer cell invasion, while expression of the kinase-mutated RAFTK-R457 (KM) or the Src binding site mutant RAFTK (402) did not affect this cell invasion. Furthermore, HRG leads to the activation of MAP kinase which is mediated by RAFTK. These findings indicate that RAFTK serves as a mediator and an integration point between the GAP proteins and HRG-mediated signaling in breast cancer cells, and implicate RAFTK involvement in the MAP kinase pathway and in breast cancer cell invasion.

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Abbreviations

aa:

amino acids

FAK:

focal adhesion kinase

GAP:

GTPase-activating protein

GST:

glutathione S-transferase

HRG:

heregulin

IP:

immunoprecipitation

RAFTK:

related adhesion focal tyrosine kinase

WB:

Western blot

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Acknowledgements

We thank Roanna London and Shuxian Jiang for their advice and technical help. We also thank Janet Delahanty for editing, Nancy DesRosiers for preparation of the figures, and Peter Park for typing of this manuscript. This work was supported in part by National Institutes of Health Grants HL55445, HL51456, DAMD 17-98-1-8032, DAMD 17-99-1-9078, and CA76226. This paper is dedicated to Charlene Engelhard and Ronald Ansin for their continuing friendship and support for our research program.

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Authors and Affiliations

  1. Division of Experimental Medicine, Beth Israel Deaconess Medical Center, Harvard Institutes of Medicine, 4 Blackfan Circle, Boston, 02115, Massachusetts, MA, USA

    Sheila Zrihan-Licht, Yigong Fu, Karin Schinkmann, Shalom Avraham & Hava Avraham

  2. Massachusetts General Hospital, Cancer Center and Harvard Medical School Building 149, 13th Street, Charlestown, 02125, Massachusetts, MA, USA

    Jeffrey Settleman

  3. Pathology Department, Beth Israel Deaconess Medical Center, 330 Brookline Avenue, Boston, 02215, Massachusetts, MA, USA

    Leslie Shaw

  4. Department of Cell Research and Immunology, Tel Aviv University, Ramat Aviv, Tel Aviv, 69978, Israel

    Iafa Keydar

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  1. Sheila Zrihan-Licht
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Zrihan-Licht, S., Fu, Y., Settleman, J. et al. RAFTK/Pyk2 tyrosine kinase mediates the association of p190 RhoGAP with RasGAP and is involved in breast cancer cell invasion. Oncogene 19, 1318–1328 (2000). https://doi.org/10.1038/sj.onc.1203422

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