Murine FATP alleviates growth and biochemical deficiencies of yeast fat1 Delta strains

Saccharomyces cerevisiae is an ideal model eukaryote for studying fatty-aci d transport. Yeast are auxotrophic for unsaturated fatty acids when grown u nder hypoxic conditions or when the fatty-acid synthase inhibitor cerulenin is included in the growth media. The FAT1 gene encodes a protein, Fat1p, w hich is required for maximal levels of fatty-acid import and has an acyl Co A synthetase activity specific for very-long-chain fatty acids suggesting t his protein plays a pivotal role in fatty-acid trafficking. In the present work, we present evidence that Fat1p and the murine fatty-acid transport pr otein (FATP) are functional homologues. FAT1 is essential for growth under hypoxic conditions and when cerulenin was included in the culture media in the presence or absence of unsaturated fatty acids. FAT1 disruptants (fat1 Delta) fail to accumulate the fluorescent long-chain fatty acid fatty-acid analogue 4,4-difluoro-5-methyl-4-bora-3a,4a-diaza-s-indacene-3-dodecanoic a cid (C-1-BODIPY-C-12), have a greatly diminished capacity to transport exog enous long-chain fatty acids, and have very long-chain acyl CoA synthetase activities that were 40% wild-type. The depression in very long-chain acyl CoA synthetase activities were not apparent in cells grown in the presence of oleate. Additionally, beta-oxidation of exogenous long-chain fatty acids is depressed to 30% wild-type levels. The reduction of beta-oxidation was correlated with a depression of intracellular oleoyl CoA levels in the fat1 Delta strain following incubation of the cells with exogenous oleate. Expr ession of either Fat1p or murine FATP from a plasmid in a fat1 Delta strain restored these phenotypic and biochemical deficiencies. Fat1p and FATP res tored growth of fat1 Delta cells in the presence of cerulenin and under hyp oxic conditions. Furthermore, fatty-acid transport was restored and was fou nd to be chain length specific: octanoate, a medium-chain fatty acid was tr ansported in a Fat1p- and FATP-independent manner while the long-chain fatt y acids myristate, palmitate, and oleate required either Fat1p or FATP for maximal levels of transport. Lignoceryl CoA synthetase activities were rest ored to wild-type levels in fat1 Delta strains expressing either Fat1p or F ATP. Fat1p or FATP also restored wild-type levels of beta-oxidation of exog enous long-chain fatty acids. These data show that Fat1p and FATP are funct ionally equivalent when expressed in yeast and play a central role in fatty -acid trafficking.