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The localization of PepT1 in intestinal brush border lipid raft microdomains determines its transport function
Author(s) -
Nguyen Hang Thi Thu,
Charrier Laetitia,
Sitaraman Shanthi,
Merlin Didier
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a588-a
Subject(s) - lipid raft , brush border , raft , ganglioside , microbiology and biotechnology , intestinal epithelium , cell fractionation , intestinal mucosa , chemistry , biophysics , biology , biochemistry , vesicle , epithelium , signal transduction , membrane , medicine , organic chemistry , copolymer , polymer , genetics
The brush border (BB) of small intestinal enterocytes is a rich source of lipid rafts, which are membrane microdomains that are enriched in cholesterol and glycosphingolipids, and contain some digestive enzymes, trafficking molecules and signaling proteins. The intestinal epithelial apical transporter, hPepT1, is expressed in intestinal epithelial cells, where it is responsible for the uptake of a broad range of small peptides. To examine the mechanism of hPepT1‐mediated peptide transport in more detail, we herein investigated the presence of hPepT1 in intestinal lipid rafts. Immunoblotting showed that hPepT1 was enriched in low‐density fractions (LDFs; containing the lipid rafts) isolated from mouse intestinal BB and polarized Caco2‐BBE cells by detergent extraction and sucrose gradient fractionation. In these fractions, hPepT1 co‐localized with N aminopeptidase, a well‐known intestinal raft resident, as well as the raft marker, ganglioside GM1. Disruption of lipid rafts by cholesterol depletion with methyl beta cyclodextrin caused hPepT1 to shift from the LDFs to the high‐density fractions and decreased the hPepT1‐mediated transport activity. Lastly, detergent‐insoluble fractions extracted from non‐polarized Caco2‐BBE cells were almost completely devoid of hPepT1. These findings collectively suggest that hPepT1 is mainly distributed in the lipid rafts from polarized enterocytes, where its activity may be lipid raft‐dependent.