Liver Fatty Acid-binding Protein Initiates Budding of Pre-chylomicron Transport Vesicles from Intestinal Endoplasmic Reticulum

Department of Chemistry, The University of Memphis, Memphis, Tennessee, United States
Journal of Biological Chemistry (Impact Factor: 4.57). 07/2007; 282(25):17974-84. DOI: 10.1074/jbc.M610765200
Source: PubMed


The rate-limiting step in the transit of absorbed dietary fat across the enterocyte is the generation of the pre-chylomicron transport vesicle (PCTV) from the endoplasmic reticulum (ER). This vesicle does not require coatomer-II (COPII) proteins for budding from the ER membrane and contains vesicle-associated membrane protein 7, found in intestinal ER, which is a unique intracellular location for this SNARE protein. We wished to identify the protein(s) responsible for budding this vesicle from ER membranes in the absence of the requirement for COPII proteins. We chromatographed rat intestinal cytosol on Sephacryl S-100 and found that PCTV budding activity appeared in the low molecular weight fractions. Additional chromatographic steps produced a single major and several minor bands on SDS-PAGE. By tandem mass spectroscopy, the bands contained both liver and intestinal fatty acid-binding proteins (L- and I-FABP) as well as four other proteins. Recombinant proteins for each of the six proteins identified were tested for PCTV budding activity; only L-FABP and I-FABP (23% the activity of L-FABP) were active. The vesicles generated by L-FABP were sealed, contained apolipoproteins B48 and AIV, were of the same size as PCTV on Sepharose CL-6B, and by electron microscopy, excluded calnexin and calreticulin but did not fuse with cis-Golgi nor did L-FABP generate COPII-dependent vesicles. Gene-disrupted L-FABP mouse cytosol had 60% the activity of wild type mouse cytosol. We conclude that L-FABP can select cargo for and bud PCTV from intestinal ER membranes.

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Available from: Charles M Mansbach, May 28, 2014
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    • "Like LFABP, IFABP is found widely distributed throughout the cytosol of the enterocyte during the fed state, but is localized toward the apical side of the cell in the fasted state [34]. IFABP has not been found to be involved with chylomicron formation [35] [57]; hence, IFABP has been proposed to be involved with uptake of FA from the lumen of the intestine, and with trafficking within the intestinal enterocyte to organelles [34]. It is also thought that both FABPs may serve as a cytosolic reservoir for FA required for various cellular functions, while also preventing accumulation of unesterified FA, which are known to modify membrane properties [4]. "
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    • "Unused protein is degraded, a mechanism that ensures that there is sufficient apo B48 for even the largest fat meal. Fat feeding increases apo AIV expression, and apo A1V serves as a surface component for apo B48 particles in the entrecote [1, 2]. Apo A IV may stimulate net transfer of membrane triglyceride to luminal particles. "
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    • "It is conceivable that the coat-independent formation of preperoxisomal vesicular carriers from the ER in mammalian cells is driven by the COPII-independent protein machine that they use for generating large prechylomicron vesicular carriers from the ER (Neeli et al., 2007; Siddiqi et al., 2003, 2006). Two components of this protein machine have been recently identified as the post-Golgi v-SNARE protein VAMP7 (Siddiqi et al., 2006) and the L-FABP (Neeli et al., 2007). We suggest that the coat-independent route of preperoxisomal vesicle formation from the ER is driven by proteins that, following their recruitment from the cytosol to the neck of the emergent bud, promote the lateral movement of specialized membrane domains containing group I PMPs and certain lipid species, thereby populating the curved portion of the membrane with these constituents of preperoxisomal vesicles, and the eventual scission of the bud neck, thus detaching preperoxisomal vesicles from the ER. "
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