Inhibition of binding to fatty acid binding protein reduces the intracellular transport of fatty acids.
Male livers, containing lesser amounts of fatty acid binding protein (FABP), utilize fatty acids more slowly than female livers. Conventional wisdom dictates that FABP stimulates fatty acid use by increasing cytoplasmic transport rates. Previously, we showed that the cytoplasmic diffusion of a fatty acid analogue [12-N-methyl-7-nitrobenzo-2-oxa-1,3-diazol-amino stearate (NBD-stearate)] is faster in female hepatocytes, paralleling the larger amounts of FABP. Sex differences in other cytoplasmic factors could also lead to faster diffusion, independent of FABP levels. The aim of this study was to determine the effect of inhibition of fatty acid binding to FABP on the directly measured intracellular transport rate of NBD-stearate. The binding of NBD-stearate to FABP was reduced by incubating hepatocytes isolated from male and female rats with alpha-bromo-palmitate (0-1,500 microM), a modified long-chain fatty acid that binds to FABP. The inhibition by alpha-bromo-palmitate on NBD-stearate binding to FABP was measured with the use of centrifugation to separate cytosol from cytoplasmic membranes. Laser photobleaching (fluorescence recovery after photobleaching) was used to measure the cytoplasmic diffusion of NBD-stearate in hepatocytes. Alpha-Bromo-palmitate incubation reduced NBD-stearate binding to FABP in a dose-dependent manner. The measured diffusion rate was also reduced in proportion to the degree of binding inhibition. We conclude that cytoplasmic transport of NBD-stearate is modulated by binding to soluble proteins like FABP. FABP enhances diffusive transport by reducing binding to immobile cytosolic membranes.
Luxon
B A
BA
Division of Gastroenterology and Hepatology, St. Louis University Health Sciences Center, Missouri 63110-0250, USA.
D009327:4-Chloro-7-nitrobenzofurazan / Q000031:analogs & derivatives / Q000378:metabolism; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004058:Diffusion; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids / Q000378:metabolism*; D005260:Female; D005453:Fluorescence; D007425:Intracellular Membranes / Q000378:metabolism*; D007834:Lasers; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008297:Male; D018996:Myelin P2 Protein / Q000037:antagonists & inhibitors* / Q000378:metabolism; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D010168:Palmitates / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012727:Sex Characteristics; D013228:Stearates / Q000378:metabolism
D002352:Carrier Proteins; C490174:Fabp7 protein, rat; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids; D018996:Myelin P2 Protein; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D010168:Palmitates; D013228:Stearates; C060027:12-N-methyl-7-nitrobenzo-2-oxa-1,3-diazolamino stearate; C022776:2-bromopalmitate; D009327:4-Chloro-7-nitrobenzofurazan