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J Cell Biol. May 1, 1995; 129(3): 619–627.
PMCID: PMC2120430

Glycolipid-anchored proteins in neuroblastoma cells form detergent- resistant complexes without caveolin

Abstract

It has been known for a number of years that glycosyl- phosphatidylinositol (GPI)-anchored proteins, in contrast to many transmembrane proteins, are insoluble at 4 degrees C in nonionic detergents such as Triton X-100. Recently, it has been proposed that this behavior reflects the incorporation of GPI-linked proteins into large aggregates that are rich in sphingolipids and cholesterol, as well as in cytoplasmic signaling molecules such as heterotrimeric G proteins and src-family tyrosine kinases. It has been suggested that these lipid-protein complexes are derived from caveolae, non-clathrin- coated invaginations of the plasmalemma that are abundant in endothelial cells, smooth muscle, and lung. Caveolin, a proposed coat protein of caveolae, has been hypothesized to be essential for formation of the complexes. To further investigate the relationship between the detergent-resistant complexes and caveolae, we have characterized the behavior of GPI-anchored proteins in lysates of N2a neuroblastoma cells, which lack morphologically identifiable caveolae, and which do not express caveolin (Shyng, S.-L., J. E. Heuser, and D. A. Harris. 1994. J. Cell Biol. 125:1239-1250). We report here that the complexes prepared from N2a cells display the large size and low buoyant density characteristic of complexes isolated from sources that are rich in caveolae, and contain the same major constituents, including multiple GPI-anchored proteins, alpha and beta subunits of heterotrimeric G proteins, and the tyrosine kinases fyn and yes. Our results argue strongly that detergent-resistant complexes are not equivalent to caveolae in all cell types, and that in neuronal cells caveolin is not essential for the integrity of these complexes.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Anderson RG. Plasmalemmal caveolae and GPI-anchored membrane proteins. Curr Opin Cell Biol. 1993 Aug;5(4):647–652. [PubMed]
  • Anderson RG. Caveolae: where incoming and outgoing messengers meet. Proc Natl Acad Sci U S A. 1993 Dec 1;90(23):10909–10913. [PMC free article] [PubMed]
  • Arreaza G, Melkonian KA, LaFevre-Bernt M, Brown DA. Triton X-100-resistant membrane complexes from cultured kidney epithelial cells contain the Src family protein tyrosine kinase p62yes. J Biol Chem. 1994 Jul 22;269(29):19123–19127. [PubMed]
  • Bohuslav J, Cinek T, Horejsí V. Large, detergent-resistant complexes containing murine antigens Thy-1 and Ly-6 and protein tyrosine kinase p56lck. Eur J Immunol. 1993 Apr;23(4):825–831. [PubMed]
  • Borchelt DR, Taraboulos A, Prusiner SB. Evidence for synthesis of scrapie prion proteins in the endocytic pathway. J Biol Chem. 1992 Aug 15;267(23):16188–16199. [PubMed]
  • Bordier C. Phase separation of integral membrane proteins in Triton X-114 solution. J Biol Chem. 1981 Feb 25;256(4):1604–1607. [PubMed]
  • Brown D. The tyrosine kinase connection: how GPI-anchored proteins activate T cells. Curr Opin Immunol. 1993 Jun;5(3):349–354. [PubMed]
  • Brown DA, Rose JK. Sorting of GPI-anchored proteins to glycolipid-enriched membrane subdomains during transport to the apical cell surface. Cell. 1992 Feb 7;68(3):533–544. [PubMed]
  • Casey PJ, Fong HK, Simon MI, Gilman AG. Gz, a guanine nucleotide-binding protein with unique biochemical properties. J Biol Chem. 1990 Feb 5;265(4):2383–2390. [PubMed]
  • Caughey B, Raymond GJ. The scrapie-associated form of PrP is made from a cell surface precursor that is both protease- and phospholipase-sensitive. J Biol Chem. 1991 Sep 25;266(27):18217–18223. [PubMed]
  • Chang WJ, Ying YS, Rothberg KG, Hooper NM, Turner AJ, Gambliel HA, De Gunzburg J, Mumby SM, Gilman AG, Anderson RG. Purification and characterization of smooth muscle cell caveolae. J Cell Biol. 1994 Jul;126(1):127–138. [PMC free article] [PubMed]
  • Cinek T, Horejsí V. The nature of large noncovalent complexes containing glycosyl-phosphatidylinositol-anchored membrane glycoproteins and protein tyrosine kinases. J Immunol. 1992 Oct 1;149(7):2262–2270. [PubMed]
  • Coyne KE, Hall SE, Thompson S, Arce MA, Kinoshita T, Fujita T, Anstee DJ, Rosse W, Lublin DM. Mapping of epitopes, glycosylation sites, and complement regulatory domains in human decay accelerating factor. J Immunol. 1992 Nov 1;149(9):2906–2913. [PubMed]
  • Cross GA. Glycolipid anchoring of plasma membrane proteins. Annu Rev Cell Biol. 1990;6:1–39. [PubMed]
  • Cullen BR. Trans-activation of human immunodeficiency virus occurs via a bimodal mechanism. Cell. 1986 Sep 26;46(7):973–982. [PubMed]
  • Englund PT. The structure and biosynthesis of glycosyl phosphatidylinositol protein anchors. Annu Rev Biochem. 1993;62:121–138. [PubMed]
  • Fra AM, Williamson E, Simons K, Parton RG. Detergent-insoluble glycolipid microdomains in lymphocytes in the absence of caveolae. J Biol Chem. 1994 Dec 9;269(49):30745–30748. [PubMed]
  • Glenney JR., Jr Tyrosine phosphorylation of a 22-kDa protein is correlated with transformation by Rous sarcoma virus. J Biol Chem. 1989 Dec 5;264(34):20163–20166. [PubMed]
  • Glenney JR., Jr The sequence of human caveolin reveals identity with VIP21, a component of transport vesicles. FEBS Lett. 1992 Dec 7;314(1):45–48. [PubMed]
  • Harris DA, Falls DL, Johnson FA, Fischbach GD. A prion-like protein from chicken brain copurifies with an acetylcholine receptor-inducing activity. Proc Natl Acad Sci U S A. 1991 Sep 1;88(17):7664–7668. [PMC free article] [PubMed]
  • Harris DA, Huber MT, van Dijken P, Shyng SL, Chait BT, Wang R. Processing of a cellular prion protein: identification of N- and C-terminal cleavage sites. Biochemistry. 1993 Feb 2;32(4):1009–1016. [PubMed]
  • Jadot M, Canfield WM, Gregory W, Kornfeld S. Characterization of the signal for rapid internalization of the bovine mannose 6-phosphate/insulin-like growth factor-II receptor. J Biol Chem. 1992 Jun 5;267(16):11069–11077. [PubMed]
  • Kurzchalia TV, Dupree P, Parton RG, Kellner R, Virta H, Lehnert M, Simons K. VIP21, a 21-kD membrane protein is an integral component of trans-Golgi-network-derived transport vesicles. J Cell Biol. 1992 Sep;118(5):1003–1014. [PMC free article] [PubMed]
  • Muirhead ML, Action RT, Williams AF. Preliminary characterization of Thy-1.1 and Ag-B antigens from rat tissues solubilized in detergents. Biochem J. 1974 Oct;143(1):51–61. [PMC free article] [PubMed]
  • Lisanti MP, Tang ZL, Sargiacomo M. Caveolin forms a hetero-oligomeric protein complex that interacts with an apical GPI-linked protein: implications for the biogenesis of caveolae. J Cell Biol. 1993 Nov;123(3):595–604. [PMC free article] [PubMed]
  • Lisanti MP, Scherer PE, Vidugiriene J, Tang Z, Hermanowski-Vosatka A, Tu YH, Cook RF, Sargiacomo M. Characterization of caveolin-rich membrane domains isolated from an endothelial-rich source: implications for human disease. J Cell Biol. 1994 Jul;126(1):111–126. [PMC free article] [PubMed]
  • Lisanti MP, Scherer PE, Tang Z, Sargiacomo M. Caveolae, caveolin and caveolin-rich membrane domains: a signalling hypothesis. Trends Cell Biol. 1994 Jul;4(7):231–235. [PubMed]
  • Low MG. The glycosyl-phosphatidylinositol anchor of membrane proteins. Biochim Biophys Acta. 1989 Dec 6;988(3):427–454. [PubMed]
  • Mayor S, Rothberg KG, Maxfield FR. Sequestration of GPI-anchored proteins in caveolae triggered by cross-linking. Science. 1994 Jun 24;264(5167):1948–1951. [PubMed]
  • Mumby SM, Kahn RA, Manning DR, Gilman AG. Antisera of designed specificity for subunits of guanine nucleotide-binding regulatory proteins. Proc Natl Acad Sci U S A. 1986 Jan;83(2):265–269. [PMC free article] [PubMed]
  • Parton RG, Joggerst B, Simons K. Regulated internalization of caveolae. J Cell Biol. 1994 Dec;127(5):1199–1215. [PMC free article] [PubMed]
  • Prusiner SB, DeArmond SJ. Prion diseases and neurodegeneration. Annu Rev Neurosci. 1994;17:311–339. [PubMed]
  • Robinson PJ. Phosphatidylinositol membrane anchors and T-cell activation. Immunol Today. 1991 Jan;12(1):35–41. [PubMed]
  • Rodgers W, Crise B, Rose JK. Signals determining protein tyrosine kinase and glycosyl-phosphatidylinositol-anchored protein targeting to a glycolipid-enriched membrane fraction. Mol Cell Biol. 1994 Aug;14(8):5384–5391. [PMC free article] [PubMed]
  • Rothberg KG, Ying YS, Kolhouse JF, Kamen BA, Anderson RG. The glycophospholipid-linked folate receptor internalizes folate without entering the clathrin-coated pit endocytic pathway. J Cell Biol. 1990 Mar;110(3):637–649. [PMC free article] [PubMed]
  • Rothberg KG, Heuser JE, Donzell WC, Ying YS, Glenney JR, Anderson RG. Caveolin, a protein component of caveolae membrane coats. Cell. 1992 Feb 21;68(4):673–682. [PubMed]
  • Sargiacomo M, Sudol M, Tang Z, Lisanti MP. Signal transducing molecules and glycosyl-phosphatidylinositol-linked proteins form a caveolin-rich insoluble complex in MDCK cells. J Cell Biol. 1993 Aug;122(4):789–807. [PMC free article] [PubMed]
  • Shenoy-Scaria AM, Dietzen DJ, Kwong J, Link DC, Lublin DM. Cysteine3 of Src family protein tyrosine kinase determines palmitoylation and localization in caveolae. J Cell Biol. 1994 Jul;126(2):353–363. [PMC free article] [PubMed]
  • Shyng SL, Huber MT, Harris DA. A prion protein cycles between the cell surface and an endocytic compartment in cultured neuroblastoma cells. J Biol Chem. 1993 Jul 25;268(21):15922–15928. [PubMed]
  • Shyng SL, Heuser JE, Harris DA. A glycolipid-anchored prion protein is endocytosed via clathrin-coated pits. J Cell Biol. 1994 Jun;125(6):1239–1250. [PMC free article] [PubMed]
  • Simons K, Wandinger-Ness A. Polarized sorting in epithelia. Cell. 1990 Jul 27;62(2):207–210. [PubMed]
  • Smart EJ, Ying YS, Conrad PA, Anderson RG. Caveolin moves from caveolae to the Golgi apparatus in response to cholesterol oxidation. J Cell Biol. 1994 Dec;127(5):1185–1197. [PMC free article] [PubMed]
  • Stefanová I, Horejsí V, Ansotegui IJ, Knapp W, Stockinger H. GPI-anchored cell-surface molecules complexed to protein tyrosine kinases. Science. 1991 Nov 15;254(5034):1016–1019. [PubMed]
  • Ulrich MJ, Ley TJ. Function of normal and mutated gamma-globin gene promoters in electroporated K562 erythroleukemia cells. Blood. 1990 Feb 15;75(4):990–999. [PubMed]
  • Vitetta ES, Boyse EA, Uhr JW. Isolation and characterization of a molecular complex containing Thy-1 antigen from the surface of murine thymocytes and T cells. Eur J Immunol. 1973 Jul;3(7):446–453. [PubMed]
  • Woodward MP, Roth TF. Coated vesicles: characterization, selective dissociation, and reassembly. Proc Natl Acad Sci U S A. 1978 Sep;75(9):4394–4398. [PMC free article] [PubMed]
  • Ying YS, Anderson RG, Rothberg KG. Each caveola contains multiple glycosyl-phosphatidylinositol-anchored membrane proteins. Cold Spring Harb Symp Quant Biol. 1992;57:593–604. [PubMed]
  • Zurzolo C, van't Hof W, van Meer G, Rodriguez-Boulan E. VIP21/caveolin, glycosphingolipid clusters and the sorting of glycosylphosphatidylinositol-anchored proteins in epithelial cells. EMBO J. 1994 Jan 1;13(1):42–53. [PMC free article] [PubMed]

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