Phospho-Caveolin-1 Mediates Integrin-Regulated Membrane Domain Internalisation

Miguel del Pozo

Nagaraj Balasubramanian

Nazilla Alderson

William Kiosses

Araceli Grande-García

Richard Anderson

Martin Schwartz

^{Correspondence should be addressed to M.A.d.P. (e-mail:}se.cinc@ozopledam)

Abstract

Growth of normal cells is anchorage-dependent because signalling through multiple pathways including Erk, PI 3-kinase and Rac requires integrin-mediated cell adhesion ¹. Components of these pathways localize to low density, cholesterol-rich domains in the plasma membrane named “lipid rafts” ²,3 or “cholesterol enriched membrane microdomains” (CEMM) ⁴. We previously reported that integrin-mediated adhesion regulates CEMM trafficking such that cell detachment from the extracellular matrix (ECM) triggers CEMM internalisation and clearance from the plasma membrane ⁵. We now report that this internalisation is mediated by dynamin-2 and caveolin-1. Internalisation requires phosphorylation of caveolin-1 on tyrosine 14. A shift in localisation of phospho-caveolin-1 from focal adhesions to caveolae induces CEMM internalisation upon cell detachment, which mediates inhibition of Erk, PI 3-kinase and Rac. These data define a novel molecular mechanism for growth and tumour suppression by caveolin-1.

Keywords: anchorage-dependent cell growth, cancer, integrin signalling, caveolin, cholesterol-enriched membrane microdomains (CEMM), Rho GTPases

Abstract

Footnotes

A significant part of this work was performed while some of the authors were located at The Scripps Research Institute, Departments of Immunology (MAdP and NBA, from January 2003 to June 2004) and Cell Biology (MAdP, NBA and WBK until June 2004, and MAS until July 2002), La Jolla, California 92037, USA.

Supplementary Information accompanies the paper on the Nature Cell Biology’s website.

Competing Interests statement The authors declare that they have no competing financial interests.

Footnotes

References

1. Schwartz MAIntegrins, oncogenes, and anchorage independence. J. Cell Biol. 1997;139:575–578.[Google Scholar]
2. Edidin MThe state of lipid rafts: from model membranes to cells. Annu Rev Biophys Biomol Struct. 2003;32:257–83.[PubMed][Google Scholar]
3. Lagerholm BC, Weinreb GE, Jacobson K, Thompson NLDetecting Microdomains in Intact Cell Membranes. Annu Rev Phys Chem. 2005;56:309–336.[PubMed][Google Scholar]
4. Grande A, Echarri A, del Pozo MAIntegrin regulation of membrane domain trafficking and Rac targeting. Biochemical Society Transactions. 2005;33:609–613.[PubMed][Google Scholar]
5. del Pozo MA, et al Integrins regulate Rac targeting by internalization of membrane domains. Science. 2004;303:839–42.[PubMed][Google Scholar]
6. del Pozo MA, Price LS, Alderson NB, Ren XD, Schwartz MAAdhesion to the extracellular matrix regulates the coupling of the small GTPase Rac to its effector PAK. Embo J. 2000;19:2008–2014.[Google Scholar]
7. Parton RGCaveolae--from ultrastructure to molecular mechanisms. Nat Rev Mol Cell Biol. 2003;4:162–7.[PubMed][Google Scholar]
8. Pelkmans L, Puntener D, Helenius ALocal actin polymerization and dynamin recruitment in SV40-induced internalization of caveolae. Science. 2002;296:535–9.[PubMed][Google Scholar]
9. Fielding CJ, Fielding PECaveolae and intracellular trafficking of cholesterol. Adv Drug Deliv Rev. 2001;49:251–64.[PubMed][Google Scholar]
10. Cao H, Courchesne WE, Mastick CCA phosphotyrosine-dependent protein interaction screen reveals a Role for phosphorylation of caveolin-1 on tyrosine 14: Recruitment of C-terminal Src kinase. J Biol Chem. 2002;22:22.[PubMed][Google Scholar]
11. Lee H, et al Constitutive and growth factor-regulated phosphorylation of caveolin-1 occurs at the same site (Tyr-14) in vivo: identification of a c-Src/Cav-1/Grb7 signaling cassette. Mol Endocrinol. 2000;14:1750–75.[PubMed][Google Scholar]
12. Engelman JA, et al Recombinant expression of caveolin-1 in oncogenically transformed cells abrogates anchorage-independent growth. J. Biol. Chem. 1997;272:16374–16381.[PubMed][Google Scholar]
13. Fiucci G, Ravid D, Reich R, Liscovitch MCaveolin-1 inhibits anchorage-independent growth, anoikis and invasiveness in MCF-7 human breast cancer cells. Oncogene. 2002;21:2365–75.[PubMed][Google Scholar]
14. Wiechen K, et al Caveolin-1 is down-regulated in human ovarian carcinoma and acts as a candidate tumor suppressor gene. Am J Pathol. 2001;159:1635–43.[Google Scholar]
15. Capozza F, et al Absence of caveolin-1 sensitizes mouse skin to carcinogen-induced epidermal hyperplasia and tumor formation. Am J Pathol. 2003;162:2029–39.[Google Scholar]
16. Williams TM, et al. Caveolin-1 gene disruption promotes mammary tumorigenesis and dramatically enhances lung metastasis in vivo. Role of Cav-1 in cell invasiveness and matrix metalloproteinase (MMP-2/9) secretion. J Biol Chem. 2004;279:51630–46.[PubMed]
17. Lee H, et al Caveolin-1 mutations (P132L and null) and the pathogenesis of breast cancer: caveolin-1 (P132L) behaves in a dominant-negative manner and caveolin-1 (-/-) null mice show mammary epithelial cell hyperplasia. Am J Pathol. 2002;161:1357–69.[Google Scholar]
18. Murata M, et al VIP21/caveolin is a cholesterol-binding protein. Proc Natl Acad Sci U S A. 1995;92:10339–43.[Google Scholar]
19. Sabharanjak S, Sharma P, Parton RG, Mayor SGPI-anchored proteins are delivered to recycling endosomes via a distinct cdc42-regulated, clathrin-independent pinocytic pathway. Dev Cell. 2002;2:411–23.[PubMed][Google Scholar]
20. Damm EM, et al Clathrin- and caveolin-1-independent endocytosis: entry of simian virus 40 into cells devoid of caveolae J Cell Biol 2005168477–88.Epub 2005 Jan 24 [Google Scholar]
21. Razani B, et al Caveolin-1 null mice are viable but show evidence of hyperproliferative and vascular abnormalities. J Biol Chem. 2001;276:38121–38.[PubMed][Google Scholar]
22. Mettouchi A, et al Integrin-specific activation of Rac controls progression through the G(1) phase of the cell cycle. Mol Cell. 2001;8:115–27.[PubMed][Google Scholar]
23. Boyd ND, Chan BM, Petersen NOBeta1 integrins are distributed in adhesion structures with fibronectin and caveolin and in coated pits. Biochem Cell Biol. 2003;81:335–48.[PubMed][Google Scholar]
24. Parton RG, Joggerst B, Simons KRegulated internalization of caveolae. J Cell Biol. 1994;127:1199–215.[Google Scholar]
25. Aoki T, Nomura R, Fujimoto TTyrosine phosphorylation of caveolin-1 in the endothelium. Exp Cell Res. 1999;253:629–36.[PubMed][Google Scholar]
26. Wary KK, Mariotti A, Zurzolo C, Giancotti FGA requirement for caveolin-1 and associated kinase Fyn in integrin signaling and anchorage-dependent cell growth. Cell. 1998;94:625–634.[PubMed][Google Scholar]
27. Palazzo AF, Eng CH, Schlaepfer DD, Marcantonio EE, Gundersen GGLocalized stabilization of microtubules by integrin- and FAK-facilitated Rho signaling. Science. 2004;303:836–9.[PubMed][Google Scholar]
28. Drab M, et al Loss of caveolae, vascular dysfunction, and pulmonary defects in caveolin-1 gene-disrupted mice. Science. 2001;293:2449–52.[PubMed][Google Scholar]
29. Sunaga N, et al Different roles for caveolin-1 in the development of non-small cell lung cancer versus small cell lung cancer. Cancer Res. 2004;64:4277–85.[PubMed][Google Scholar]
30. Machleidt T, Li WP, Liu P, Anderson RGMultiple domains in caveolin-1 control its intracellular traffic. J. Cell Biol. 2000;148:17–28.[Google Scholar]