A Novel Type of Detergent-resistant Membranes May Contribute to an Early Protein Sorting Event in Epithelial Cells

• Published in: The Journal of biological chemistry 2005-12, Vol.280 (52), p.42636-42643
• Main Authors: Alfalah, Marwan, Wetzel, Gabi, Fischer, Ina, Busche, Roger, Sterchi, Erwin E., Zimmer, Klaus-Peter, Sallmann, Hans-Peter, Naim, Hassan Y.
• Format: Article
• Language: English
• Subjects: Adenosine Deaminase - chemistry; Animals; Caco-2 Cells; CD13 Antigens - pharmacology; Cell Line; Cell Membrane - metabolism; Chlorocebus aethiops; Cholesterol - metabolism; COS Cells; Detergents - pharmacology; Dipeptidyl Peptidase 4 - chemistry; Dogs; Dose-Response Relationship, Drug; Drug Resistance; Endoplasmic Reticulum - metabolism; Epithelial Cells - cytology; Epithelial Cells - pathology; Glycoproteins - chemistry; Golgi Apparatus - metabolism; Humans; Hybridomas - metabolism; Immunoprecipitation; Kinetics; Lactase - chemistry; Lactase-Phlorizin Hydrolase - chemistry; Lipids - chemistry; Mannose - chemistry; Membrane Cofactor Protein - biosynthesis; Membrane Microdomains - chemistry; Mice; Octoxynol - pharmacology; Oligo-1,6-Glucosidase - chemistry; Phosphatidylglycerols - chemistry; Phosphatidylinositols - chemistry; Polysorbates - pharmacology; Protein Transport; Sphingolipids - metabolism; Sucrase - chemistry; Sucrose - pharmacology; trans-Golgi Network - metabolism; Transfection; Vesicular stomatitis virus
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M505924200

One sorting mechanism of apical and basolateral proteins in epithelial cells is based on their solubility profiles with Triton X-100. Nevertheless, apical proteins themselves are also segregated beyond the trans-Golgi network by virtue of their association or nonassociation with cholesterol/sphingolipid-rich microdomains (Jacob, R., and Naim, H. Y. (2001) Curr. Biol. 11, 1444–1450). Therefore, extractability with Triton X-100 does not constitute an absolute criterion of protein sorting. Here, we investigate the solubility patterns of apical and basolateral proteins with other detergents and demonstrate that the mild detergent Tween 20 is adequate to discriminate between apical and basolateral proteins during early stages in their biosynthesis. Although the mannose-rich forms of the apical proteins sucrase-isomaltase, lactase-phlorizin hydrolase, aminopeptidase N, and dipeptidylpeptidase IV reveal similar solubility profiles comprising soluble and nonsoluble fractions, the basolateral proteins, vesicular stomatitis virus G protein, major histocompatibility complex class I, and CD46 are entirely soluble with this detergent. The insoluble Tween 20 membranes are enriched in phosphatidylinositol and phosphatidylglycerol compatible with their synthesis in the endoplasmic reticulum and the existence of a novel class of detergent-resistant membranes. The association of the mannose-rich biosynthetic forms of the apical proteins, sucraseisomaltase, lactase-phlorizin hydrolase, aminopeptidase N, and dipeptidylpeptidase IV with the Tween 20-resistant membranes suggests an early polarized sorting mechanism prior to maturation in the Golgi apparatus.