title: Liver fatty acid-binding protein initiates budding of pre-chylomicron transport vesicles from intestinal endoplasmic reticulum.
abstract: 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.
authors: Department of Molecular Sciences, University of Tennessee Health Science Center, Memphis, Tennessee 38163.
authors: Indira
authors: I
authors: Neeli
authors: Division of Gastroenterology, University of Tennessee Health Science Center, Memphis, Tennessee 38163.
authors: Shadab A
authors: SA
authors: Siddiqi
authors: Division of Gastroenterology, University of Tennessee Health Science Center, Memphis, Tennessee 38163.
authors: Shahzad
authors: S
authors: Siddiqi
authors: Veterans Affairs Medical Center, Memphis, Tennessee 38104.
authors: James
authors: J
authors: Mahan
authors: Department of Nutritional Sciences, Rutgers University, New Brunswick, New Jersey 08901.
authors: William S
authors: WS
authors: Lagakos
authors: Department of Pathobiology, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, Texas 77843.
authors: Bert
authors: B
authors: Binas
authors: Department of Chemistry, University of Memphis, Memphis, Tennessee 38152.
authors: Tarun
authors: T
authors: Gheyi
authors: Department of Nutritional Sciences, Rutgers University, New Brunswick, New Jersey 08901. Electronic address: storch@aesop.rutgers.edu.
authors: Judith
authors: J
authors: Storch
authors: Division of Gastroenterology, University of Tennessee Health Science Center, Memphis, Tennessee 38163; Veterans Affairs Medical Center, Memphis, Tennessee 38104. Electronic address: cmansbach@utmem.edu.
authors: Charles M
authors: CM
authors: Mansbach
mesh_terms: D000180:Acrylic Resins / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport; D002914:Chylomicrons / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D050556:Fatty Acid-Binding Proteins / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D008099:Liver / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D000180:Acrylic Resins; D002914:Chylomicrons; D050556:Fatty Acid-Binding Proteins; D011994:Recombinant Proteins