M-CSF Regulates the Cytoskeleton via Recruitment of a Multimeric Signaling Complex to c-Fms Tyr-559/697/721*
- Roberta Faccioद,
- Sunao Takeshita‡∥,
- Graziana Colaianni‡¶,
- Jean Chappel‡,
- Alberta Zallone¶,
- Steven L. Teitelbaum‡ and
- F. Patrick Ross‡1
- Departments of ‡Pathology and Immunology and §Orthopaedic Surgery, Washington University School of Medicine, St. Louis, Missouri 63110, ¶Department of Human Anatomy, School of Medicine, University of Bari, Policlinico 70100 Bari, Italy, and ∥Department of Bone and Joint Disease, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, Aichi 474-8522, Japan
- 1 To whom correspondence should be addressed: Washington University School of Medicine, Dept. of Pathology and Immunology, Campus Box 8118, 660 South Euclid Ave., St. Louis, MO 63110. Tel.: 314-454-8079; Fax: 314-454-5505; E-mail: rossf{at}wustl.edu.
Abstract
M-CSF is known to induce cytoskeletal reorganization in macrophages and osteoclasts by activation of phosphatidylinositol 3-kinase (PI3K) and c-Src, but the detailed mechanisms remain unclear. We find, unexpectedly, that tyrosine (Tyr) to phenylalanine (Phe) mutation of Tyr-721, the PI3K binding site in the M-CSF receptor c-Fms, fails to suppress cytoskeletal remodeling or actin ring formation. In contrast, mutation of c-Fms Tyr-559 to Phe blocks M-CSF-induced cytoskeletal reorganization by inhibiting formation of a Src Family Kinase SFK·c-Cbl·PI3K complex and the downstream activation of Vav3 and Rac, two key mediators of actin remodeling. Using an add-back approach in which specific Tyr residues are reinserted into c-Fms inactivated by the absence of all seven functionally important Tyr residues, we find that Tyr-559 is necessary but not sufficient to transduce M-CSF-dependent cytoskeletal reorganization. Furthermore, this same add-back approach identifies important roles for Tyr-697 and Tyr-721 in collaborating with Tyr-559 to recruit a multimeric signaling complex that can transduce signals from c-Fms to the actin cytoskeleton.
Footnotes
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↵2 The abbreviations used are: OC, osteoclast; RTK, receptor tyrosine kinase; WT, wild type; SFK, Src family kinase; PI3K, phosphatidylinositol 3-kinase; Epo, erythropoietin; EpoR, Epo receptor; RANKL, receptor activator of NFκB ligand; BMM, bone marrow-derived macrophage; Ab, antibody; ERK, extracellular signal-regulated kinase.
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↵* This study was supported by National Institutes of Health Grants AR32788, AR46523, and AR48853 (to S. L. T.), AR52921 (to R. F.), and AR46852 and AR48812 (to F. P. R.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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- Received November 28, 2006.
- Revision received February 23, 2007.
