Microvillar Membrane Microdomains Exist at Physiological Temperature

Lipid rafts (glycosphingolipid/cholesterol-enriched membrane microdomains) have been isolated as low temperature, detergent-resistant membranes from many cell types, but despite their presumed importance as lateral sorting and signaling platforms, fundamental questions persist concerning raft function and even existence in vivo. The nonionic detergent Brij 98 was used to isolate lipid rafts from microvillar membrane vesicles of intestinal brush borders at physiological temperature to compare with rafts, obtained by “conventional” extraction using Triton X-100 at low temperature. Microvillar rafts prepared by the two protocols were morphologically different but had essentially similar profiles of protein- and lipid components, showing that raft microdomains do exist at 37 °C and are not “low temperature artifacts. ” We also employed a novel method of sequential detergent extraction at increasing temperature to define a fraction of highly detergent-resistant “superrafts. ” These were enriched in galectin-4, a β-galactoside-recognizing lectin residing on the extracellular side of the membrane. Superrafts also harbored the glycosylphosphatidylinositol-linked alkaline phosphatase and the transmembrane aminopeptidase N, whereas the peripheral lipid raft protein annexin 2 was essentially absent. In conclusion, in the microvillar membrane, galectin-4, functions as a core raft stabilizer/organizer for other, more loosely raft-associated proteins. The superraft analysis might be applicable to other membrane microdomain systems. Lipid rafts (glycosphingolipid/cholesterol-enriched membrane microdomains) have been isolated as low temperature, detergent-resistant membranes from many cell types, but despite their presumed importance as lateral sorting and signaling platforms, fundamental questions persist concerning raft function and even existence in vivo. The nonionic detergent Brij 98 was used to isolate lipid rafts from microvillar membrane vesicles of intestinal brush borders at physiological temperature to compare with rafts, obtained by “conventional” extraction using Triton X-100 at low temperature. Microvillar rafts prepared by the two protocols were morphologically different but had essentially similar profiles of protein- and lipid components, showing that raft microdomains do exist at 37 °C and are not “low temperature artifacts. ” We also employed a novel method of sequential detergent extraction at increasing temperature to define a fraction of highly detergent-resistant “superrafts. ” These were enriched in galectin-4, a β-galactoside-recognizing lectin residing on the extracellular side of the membrane. Superrafts also harbored the glycosylphosphatidylinositol-linked alkaline phosphatase and the transmembrane aminopeptidase N, whereas the peripheral lipid raft protein annexin 2 was essentially absent. In conclusion, in the microvillar membrane, galectin-4, functions as a core raft stabilizer/organizer for other, more loosely raft-associated proteins. The superraft analysis might be applicable to other membrane microdomain systems. Lipid rafts are ordered (lo phase) membrane microdomains consisting mainly of glycosphingolipids and cholesterol in the outer leaflet of the bilayer that serve as lateral platforms for many types of proteins in the cell membrane (1Simons K. Ikonen E. Nature. 1997; 387: 569-572Crossref PubMed Scopus (8188) Google Scholar, 2Brown D. A. London E. J. Biol. Chem. 2000; 275: 17221-17224Abstract Full Text Full Text PDF PubMed Scopus (2073) Google Scholar, 3Hooper N. M. Mol. Membr. Biol. 1999; 16: 145-156Crossref PubMed Scopus (364) Google Scholar). The lipid raft hypothesis was originally proposed to explain how lipids and proteins are sorted to the apical surface of polarized cells (4Simons K. Van Meer G. Biochemistry. 1988; 27: 6197-6202Crossref PubMed Scopus (1095) Google Scholar), but in recent years the raft concept has expanded into other areas of biology, including signal transduction (5Simons K. Toomre D. Nat. Rev. Mol. Cell. Biol. 2000; 1: 31-39Crossref PubMed Scopus (5207) Google Scholar, 6Bickel P. E. Am. J. Physiol. 2002; 282: E1-E10Crossref PubMed Google Scholar, 7Werlen G. Palmer E. Curr. Opin. Immunol. 2002; 14: 299-305Crossref PubMed Scopus (114) Google Scholar), pathogen invasion (8Campbell S. M. Crowe S. M. Mak J. J. Clin. Virol. 2001; 22: 217-227Crossref PubMed Scopus (233) Google Scholar, 9van der Goot F. G. Harder T. Semin. Immunol. 2001; 13: 89-97Crossref PubMed Scopus (228) Google Scholar, 10Shin J. S. Abraham S. N. Science. 2001; 293: 1447-1448Crossref PubMed Scopus (70) Google Scholar), cholesterol homeostasis and transport (11Krieger M. Annu. Rev. Biochem. 1999; 68: 523-558Crossref PubMed Scopus (461) Google Scholar, 12Simons K. Ikonen E. Science. 2000; 290: 1721-1726Crossref PubMed Scopus (1082) Google Scholar), drug targeting (13McIntosh D. P. Tan X. Y. Oh P. Schnitzer J. E. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 1996-2001Crossref PubMed Scopus (182) Google Scholar), and biomedicine (14Simons K. Ehehalt R. J. Clin. Invest. 2002; 110: 597-603Crossref PubMed Scopus (937) Google Scholar). However, despite their popularity, fundamental questions concerning the functional relevance of lipid rafts in vivo persist (15Jacobson K. Dietrich C. Trends Cell Biol. 1999; 9: 87-91Abstract Full Text Full Text PDF PubMed Scopus (378) Google Scholar, 16Edidin M. Science's STKE. 2001; (http: //stke. sciencemag. org/cgi/content/full/OCₛigtrans;2001/67/pe1) PubMed Google Scholar). To complicate matters further, recent work has made it increasingly clear that the most commonly used experimental criterion for lipid rafts, detergent insolubility at low temperature combined with flotation in a density gradient (17Brown D. A. Rose J. K. Cell. 1992; 68: 533-544Abstract Full Text PDF PubMed Scopus (2625) Google Scholar), may define different subsets of rafts, depending on the type of detergent used (18Roper K. Corbeil D. Huttner W. B. Nat. Cell Biol. 2000; 2: 582-592Crossref PubMed Scopus (483) Google Scholar, 19Taylor C. M. Coetzee T. Pfeiffer S. E. J. Neurochem. 2002; 81: 993-1004Crossref PubMed Scopus (104) Google Scholar, 20Drevot P. Langlet C. Guo X. J. Bernard A. M. Colard O. Chauvin J. P. Lasserre R. He H. T. EMBO J. 2002; 21: 1899-1908Crossref PubMed Scopus (279) Google Scholar, 21Gomez-Mouton C. Abad J. L. Mira E. Lacalle R. A. Gallardo E. Jimenez-Baranda S. Illa I. Bernad A. Manes S. Martinez A. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 9642-9647Crossref PubMed Scopus (439) Google Scholar). A common current view considers lipid rafts as small, dynamic assemblies of lipids and proteins with the ability to cluster into larger ordered platforms that may serve in specialized, cell type-dependent functions, such as nutrient absorption, cell-cell communication, or endocytosis (2Brown D. A. London E. J. Biol. Chem. 2000; 275: 17221-17224Abstract Full Text Full Text PDF PubMed Scopus (2073) Google Scholar, 14Simons K. Ehehalt R. J. Clin. Invest. 2002; 110: 597-603Crossref PubMed Scopus (937) Google Scholar, 22Anderson R. G. Jacobson K. Science. 2002; 296: 1821-1825Crossref PubMed Scopus (1013) Google Scholar, 23Edidin M. Trends Cell Biol. 2001; 11: 492-496Abstract Full Text Full Text PDF PubMed Scopus (225) Google Scholar). These developments have turned the raft concept increasingly complex, and underline the fact that much is still unknown about lipid rafts concerning their molecular organization, dynamics of assembly/disassembly, and mechanism of function. The brush border membrane of small intestinal enterocytes is a highly specialized cell surface involved in a multitude of digestive and absorptive functions (24Trier J. S. Code C. F. Handbook of Physiology: Alimentary Canal. 6. American Physiological Society, Washington, D. C. 1968: 1125-1176Google Scholar). The microvilli of this membrane are particularly rich in glycosphingolipids and cholesterol (25Christiansen K. Carlsen J. Biochim. Biophys. Acta. 1981; 647: 188-195Crossref PubMed Scopus (88) Google Scholar), and several proteins, including some of the major digestive enzymes, such as aminopeptidase N and sucrase-isomaltase (26Danielsen E. M. Biochemistry. 1995; 34: 1596-1605Crossref PubMed Scopus (129) Google Scholar, 27Mirre C. Monlauzeur L. Garcia M. Delgrossi M. H. Le Bivic A. Am. J. Physiol. 1996; 271: C887-C894Crossref PubMed Google Scholar, 28Alfalah M. Jacob R. Preuss U. Zimmer K. P. Naim H. Naim H. Y. Curr. Biol. 1999; 9: 593-596Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar), and galectin-4 (29Danielsen E. M. van Deurs B. Mol. Biol. Cell. 1997; 8: 2241-2251Crossref PubMed Scopus (90) Google Scholar) have been shown to reside in Triton X-100-insoluble lipid rafts. Microvillar rafts may well be unique in their molecular organization, because unlike most other types of lipid raft membranes, such as caveolae (30Anderson R. G. Annu. Rev. Biochem. 1998; 67: 199-225Crossref PubMed Scopus (1733) Google Scholar), those prepared from microvilli are cholesterol-independent, and caveolin-1, a commonly used lipid raft marker in other cell types, is essentially excluded from microvillar rafts (31Hansen G. H. Immerdal L. Thorsen E. Niels-Christiansen L. L. Nystrom B. T. Demant E. J. Danielsen E. M. J. Biol. Chem. 2001; 276: 32338-32344Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar). In the present work, we aimed first to answer the fundamental question whether microvillar rafts are purely a “low temperature” phenomenon, created by the conditions normally used for detergent extraction. To tackle this problem, microvillar rafts were prepared by using Brij 98, a detergent recently described for isolation of the T cell receptor signaling machinery at a physiological temperature (20Drevot P. Langlet C. Guo X. J. Bernard A. M. Colard O. Chauvin J. P. Lasserre R. He H. T. EMBO J. 2002; 21: 1899-1908Crossref PubMed Scopus (279) Google Scholar). Second, we studied in closer detail the molecular organization of lipid rafts prepared from microvillar membrane vesicles by sequential extraction with Triton X-100 at increasing temperature. Here, a fraction of “superrafts” was isolated and characterized with galectin-4 as the predominant protein. Altogether, our data show that microvillar rafts do exist also at a physiological temperature. In addition, the results underline the importance of galectin-4 functioning as a central organizer/stabilizer of lipid rafts in the microvillar membrane with other classes of membrane proteins depending, at least partially, on this lectin for their association with these microdomains. Rabbit polyclonal antibodies to galectin-4, lactase, and aminopeptidase N were those described previously (29Danielsen E. M. van Deurs B. Mol. Biol. Cell. 1997; 8: 2241-2251Crossref PubMed Scopus (90) Google Scholar, 32Hansen G. H. Sjostrom H. Noren O. Dabelsteen E. Eur. J. Cell Biol. 1987; 43: 253-259PubMed Google Scholar, 33Skovbjerg H. Noren O. Sjostrom H. Danielsen E. M. Enevoldsen B. S. Biochim. Biophys. Acta. 1982; 707: 89-97Crossref PubMed Scopus (40) Google Scholar). A rabbit antibody to alkaline phosphatase was from Biogenesis (Poole, UK), and a mouse monoclonal antibody to annexin 2 was obtained from Transduction Laboratories (Lexington, KY). Horseradish peroxidase-coupled swine anti-rabbit IgG and rabbit anti-mouse IgG were from DAKO (Glostrup, Denmark). Gold-labeled goat anti-rabbit immunoglobulin was from Amersham Biosciences. Methyl-β-cyclodextrin, Brij 98 (polyoxyethylene 20 oleyl ether), and lipid standards were purchased from Sigma, and DynabeadsTM M-500 Subcellular was from Dynal (Oslo, Norway). Pig small intestines were kindly given by Letty Klarskov and Mette Olesen (Department of Experimental Medicine, The Panum Institute, Copenhagen, Denmark). Right-side-out microvillar vesicles were prepared from small intestinal mucosa by the divalent cation precipitation technique (34Booth A. G. Kenny A. J. Biochem. J. PubMed Scopus Google Scholar). were in 2 and using a The was by at and was to a of at on the was at for to and the was at for to a of microvillar membrane common was used in lipid raft with the two nonionic Brij 98 and Triton X-100 as well as in of A lipid raft analysis by detergent extraction by gradient (17Brown D. A. Rose J. K. Cell. 1992; 68: 533-544Abstract Full Text PDF PubMed Scopus (2625) Google Scholar) was essentially as previously described (26Danielsen E. M. Biochemistry. 1995; 34: 1596-1605Crossref PubMed Scopus (129) Google Scholar), with the that the was in a of with a gradient of on with Triton X-100 and with Brij 98 was on and at 37 of detergent in were to the microvillar membrane vesicles to a detergent of the were in for analysis by and the were by a with and at to a of lipid rafts for and Lipid rafts were prepared by extraction with Triton X-100 by density gradient as described in the of the gradient were in and with Triton X-100 for at temperature. The was at for and the was in the and with Triton X-100 for at 37 at for the was lipid analysis of membranes, microvillar vesicles and lipid rafts were with (25Christiansen K. Carlsen J. Biochim. Biophys. Acta. 1981; 647: 188-195Crossref PubMed Scopus (88) Google K. J. Neurochem. PubMed Scopus Google Scholar) as previously described (31Hansen G. H. Immerdal L. Thorsen E. Niels-Christiansen L. L. Nystrom B. T. Demant E. J. Danielsen E. M. J. Biol. Chem. 2001; 276: 32338-32344Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar). of the lipid of were to analysis with lipid standards on was from other lipids in and were in cholesterol was with a and with a in was to Nature. PubMed Scopus Google Scholar). and of proteins was with antibodies to galectin-4 (29Danielsen E. M. van Deurs B. Mol. Biol. Cell. 1997; 8: 2241-2251Crossref PubMed Scopus (90) Google Scholar), H. Noren O. Sjostrom H. Danielsen E. M. Enevoldsen B. S. Biochim. Biophys. Acta. 1982; 707: 89-97Crossref PubMed Scopus (40) Google Scholar), aminopeptidase N, alkaline and annexin were by to the by the Superrafts and lipid rafts, prepared by extraction with Triton X-100 on or Brij 98 at 37 were isolated on with antibodies to galectin-4 and aminopeptidase N, by a previously described (31Hansen G. H. Immerdal L. Thorsen E. Niels-Christiansen L. L. Nystrom B. T. Demant E. J. Danielsen E. M. J. Biol. Chem. 2001; 276: 32338-32344Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar). Lipid rafts, isolated from microvillar membrane vesicles using Triton X-100 or Brij 98, were in in for 2 at a in the lipid rafts were with in for at in of and in The and lipid rafts isolated on were in with and in as described with the that the and was in were on in in and and in a with a using antibody aminopeptidase N was on of lipid rafts and as previously described G. H. L. L. Sjostrom H. Noren O. J. 1992; PubMed Scopus Google Scholar). The brush border of small intestinal enterocytes is well for on the molecular organization of a specialized cell membrane. microvilli a of vesicles that are isolated from other cell membranes by the divalent cation precipitation technique (34Booth A. G. Kenny A. J. Biochem. J. PubMed Scopus Google Scholar). We have previously used flotation in a density gradient Triton X-100 extraction on for isolation of lipid rafts and that several microvillar proteins into the rafts. the alkaline phosphatase is and the transmembrane aminopeptidase N and sucrase-isomaltase whereas is essentially excluded from microvillar rafts (26Danielsen E. M. Biochemistry. 1995; 34: 1596-1605Crossref PubMed Scopus (129) Google Scholar). In addition, galectin-4, a of the of proteins K. Annu. Rev. Cell Biol. 9: PubMed Scopus Google Scholar, H. J. Biol. Chem. Full Text PDF PubMed Google Scholar), has been as a microvillar protein that in rafts and with some of the brush border (29Danielsen E. M. van Deurs B. Mol. Biol. Cell. 1997; 8: 2241-2251Crossref PubMed Scopus (90) Google Scholar). In the present work, these microvillar proteins were used as for lipid rafts, prepared using Triton X-100 or Brij a of the flotation in a density gradient of microvillar membrane proteins by Triton X-100 on or Brij 98 at 37 subsets of proteins into the of their association with lipid rafts. of the profiles of lipid raft-associated proteins isolated with the two many common but some in the of several were In addition, the of the rafts rafts with Triton X-100 were and of low whereas those with Brij 98 were more in The in the density gradient of the microvillar proteins described was by shown in 2 galectin-4 was present in the raft of Triton microvillar membranes and also raft-associated in the Brij 98 to galectin-4, essentially was in the of the gradient that proteins, of the detergent used for the membrane extraction 2 a marker microvillar proteins. The alkaline phosphatase was present in the lipid raft with 2 whereas the transmembrane aminopeptidase N was raft-associated with 2 The lipids of microvillar rafts, isolated extraction with the two were by rafts, prepared using Triton have been shown to cholesterol and to be particularly enriched in glycosphingolipids (31Hansen G. H. Immerdal L. Thorsen E. Niels-Christiansen L. L. Nystrom B. T. Demant E. J. Danielsen E. M. J. Biol. Chem. 2001; 276: 32338-32344Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar). of raft lipids were in by the lipid profiles of those isolated by of Brij 98 and Triton X-100 were essentially that the of the two were However, rafts obtained with Brij 98 of the with Brij 98 that microvillar lipid raft microdomains isolated at a physiological temperature were similar to those prepared using Triton X-100 at low temperature. it be that microvillar rafts are not “low temperature artifacts. ” shown in lipid rafts, prepared as described had a different depending on the type of detergent used for extraction. isolated with Triton X-100 as a of with in the of In addition, some more of vesicles were aminopeptidase N was the was also in microvilli as well as microvillar membrane vesicles (31Hansen G. H. Immerdal L. Thorsen E. Niels-Christiansen L. L. Nystrom B. T. Demant E. J. Danielsen E. M. J. Biol. Chem. 2001; 276: 32338-32344Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar), but in some of the more was also the raft a bilayer Lipid rafts isolated using Brij 98 as detergent were as membrane with of but some were also present N was in membranes of the vesicles and the In conclusion, Triton X-100 extraction on in of whereas extraction with Brij 98 at 37 °C the microvillar vesicles into membrane The more of the the in lipid raft density by the results that raft is to the small with to protein and lipid We have previously that microvillar rafts, isolated by extraction with Triton are cholesterol-independent, because for caveolae in other cell types, are to the membranes with to the of detergent (31Hansen G. H. Immerdal L. Thorsen E. Niels-Christiansen L. L. Nystrom B. T. Demant E. J. Danielsen E. M. J. Biol. Chem. 2001; 276: 32338-32344Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar). To whether this was microvillar membrane vesicles were by with (31Hansen G. H. Immerdal L. Thorsen E. Niels-Christiansen L. L. Nystrom B. T. Demant E. J. Danielsen E. M. J. Biol. Chem. 2001; 276: 32338-32344Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar, G. H. Niels-Christiansen L. L. Thorsen E. Immerdal L. Danielsen E. M. J. Biol. Chem. 2000; 275: Full Text Full Text PDF PubMed Scopus (140) Google Scholar) extraction with Brij 98 at 37 of the microvillar cholesterol is by this (31Hansen G. H. Immerdal L. Thorsen E. Niels-Christiansen L. L. Nystrom B. T. Demant E. J. Danielsen E. M. J. Biol. Chem. 2001; 276: 32338-32344Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar), but as shown in a of in the was obtained to that with Brij 98 small of aminopeptidase N, alkaline and galectin-4 were to the protein. it be that the cholesterol of microvillar rafts a membrane microdomain organization that at physiological temperature and is not a that be to low temperature extraction with Triton of lipid rafts using Triton it is to the extraction at low temperature to the and these because at Triton X-100 lipids and proteins of microvillar membrane vesicles not However, microvillar lipid rafts, prepared by Triton X-100 extraction on were with the first at 20 °C and at 37 a fraction of the lipid raft membranes in Triton X-100 even at the temperature A of these temperature Triton X-100-insoluble membranes with the lipid raft from were that are enriched in a small of proteins. A analysis by a of the different microvillar lipid raft proteins with to their into the the most protein of the was galectin-4, extraction with Triton at 20 and 37 Second, the alkaline phosphatase was at 20 and 37 °C but detergent extraction. the major of the transmembrane protein aminopeptidase N was but a fraction of this microvillar protein with the most of the lipid raft-associated annexin a peripheral membrane was by Triton X-100 at 20 and small were present in the superraft The lipid of a similar to microvillar membranes and rafts, isolated by Triton X-100 or Brij 98 However, the of to a to was larger for the other two raft microvillar membranes from enterocytes are rich in glycosphingolipids of membrane that more the of lipid in the leaflet (25Christiansen K. Carlsen J. Biochim. Biophys. Acta. 1981; 647: 188-195Crossref PubMed Scopus (88) Google Scholar). The for the that in the outer leaflet of the membranes the sequential at increasing temperature. The in the superraft fraction most the leaflet of the core of these lipid rafts. had a well membrane consisting of of about in were in the and with N antibody that the and In with lipid rafts isolated by extraction with Triton X-100 the of was more in with the that a fraction of aminopeptidase N was from the membranes of the the described be membrane created by the sequential detergent extraction at increasing temperature, the used for their a method for the of the molecular that exist the lipid and protein that define lipid rafts. this the protein of that galectin-4 is the major raft the proteins of the microvillar membrane. To this were with Triton X-100 at 37 °C a in the or of a for shown in the of the of but not galectin-4 from the that at least some of this lectin is raft-associated However, in to galectin-4, of alkaline phosphatase and aminopeptidase N were also by that their raft association is at least also galectin-4, alkaline and aminopeptidase N from lipid rafts, isolated by Brij and even from microvillar membranes, with detergent showing that the association these microvillar proteins is not by detergent extraction. the results obtained with the that galectin-4 be characterized as organizer/stabilizer microvillar lipid rafts. the lectin the core of a the alkaline phosphatase and the transmembrane aminopeptidase N may more peripheral raft proteins with for these microdomains. the of the lipid raft concept into many of biology, fundamental questions concerning the functional relevance of these microdomains in vivo To a this has to do with the most used criterion for raft detergent insolubility at low temperature. given the of membrane lipids (2Brown D. A. London E. J. Biol. Chem. 2000; 275: 17221-17224Abstract Full Text Full Text PDF PubMed Scopus (2073) Google Scholar), lipid rafts well be a to the commonly used method for their this problem, Brij 98, a of the of nonionic was recently used for of lipid rafts at 37 °C from T cells (20Drevot P. Langlet C. Guo X. J. Bernard A. M. Colard O. Chauvin J. P. Lasserre R. He H. T. EMBO J. 2002; 21: 1899-1908Crossref PubMed Scopus (279) Google Scholar). These rafts had a lipid of lipid rafts and harbored the of a functional T cell receptor In the present work, microvillar proteins were used as of had a lipid raft using Triton X-100 and with to type of membrane Brij 98 extraction at 37 °C microvillar rafts a marker protein essentially similar to lipid rafts, isolated by of Triton X-100 at low temperature. The fact that Brij 98 at 37 °C lipid rafts from two different membrane as T cell membranes and microvillar membrane vesicles to the existence and functional relevance of these microdomains. the Triton characterized in this work are membranes created by the sequential detergent extraction at increasing temperature, but the of galectin-4 to as a organizer/stabilizer of these membrane microdomains. other of the galectin-4 is a protein a signal for membrane (29Danielsen E. M. van Deurs B. Mol. Biol. Cell. 1997; 8: 2241-2251Crossref PubMed Scopus (90) Google Scholar, H. J. Biol. Chem. Full Text PDF PubMed Google Scholar). that galectin-4 is by a present on microvillar membrane lipids as well as are to serve as for this lectin and it to detergent-resistant The of microvillar raft on may explain their to cholesterol have been described in the and of on the surface of T cells by to C. J. Immunol. 1999; PubMed Google Scholar). the microvillar of digestive may to these at the cell surface by from into the of the and also from function may be because the intestinal is a and many microvillar are to from the membrane by the of We these microvillar rafts as and microdomains (31Hansen G. H. Immerdal L. Thorsen E. Niels-Christiansen L. L. Nystrom B. T. Demant E. J. Danielsen E. M. J. Biol. Chem. 2001; 276: 32338-32344Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar), to the current view that lipid rafts are and small M. Trends Cell Biol. 2001; 11: 492-496Abstract Full Text Full Text PDF PubMed Scopus (225) Google Scholar). In a recently proposed the molecular for proteins to rafts is a lipid R. G. Jacobson K. Science. 2002; 296: 1821-1825Crossref PubMed Scopus (1013) Google Scholar). these have of and about lipid that as a in the lipid bilayer for the protein raft such as in a by of and applicable to other, more dynamic such as of signal transduction the lipid hypothesis the molecular organization of microvillar rafts. We that the lipid raft concept to be to the of membrane functions that on microdomains. In this we the for be applicable to other raft membrane and might be a for the of molecular these microdomains.