Purpose of review The 11 long-chain (ACSL) and very long chain

Purpose of review The 11 long-chain (ACSL) and very long chain acyl-coenzyme A (acyl-CoA) synthetases [(ACSVL)/fatty acid transport protein] are receiving considerable attention because it has become apparent that their individual functions are not redundant. pathways. An unsettled controversy focuses on the ACSVL isoforms and whether these have both enzymatic and transport functions. Another issue is whether conversion of a fatty acid to an acyl-CoA produces an increase in the AMP/ATP ratio that is sufficient to activate AMP-activated kinase. Summary FuturestudiesarerequiredtodeterminethesubcellularlocationofeachACSLandACSVL isoform and the functional importance of phosphorylation and acetylation. Purification and crystallization of mammalian ACSL and ACSVL isoforms is needed to confirm the mechanism of action and discover how these enzymesdiffer in their affinity for fatty acids of differentchainlengths.Functionally itwillbeimportanttolearnhowtheACSLisoformscan direct their acyl-CoA products toward independent downstream pathways. liver. In contrast primary epididymal adipocytes HIP from ACSL1 CH5424802 adipose-specific knockout mice have unchanged FA incorporation into TAG diacylglycerol (DAG) or phospholipid (PL) but reduced β-oxidation products (J.M. Ellis R.A. Coleman unpublished data). However siRNA knockdown of ACSL1 in 3T3-L1 adipocytes does not affect de-novo lipogenesis or FA oxidation but increases FA efflux [13??] leading the authors to conclude that ACSL1 activates FA that is re-esterified after lipolysis. Pulse-chase labeling experiments could verify reduced rates of re-esterification and confirm this interpretation. Knocking down ACSL3 in rat primary hepatocytes decreases labeled acetate incorporation into TAG and PL [14??]. In human hepatoma Huh7 cells siRNA knockdown of ACSL3 decreases oleate incorporation into phosphatidylcholine (PC) and secreted VLDL and increases cellular-free oleate whereas knockdown of ACSL1 and ACSL4 does not alter oleate incorporation into cholesterol ester (CE) TAG phospholipids or free FA [15??]. Differential effects of individual ACSL iso-forms support the notion that ACSL isoforms channel FA to distinct metabolic fates. Several overexpression studies are inconsistent with the knockdown studies and provide strikingly contradictory interpretations of the role of ACSL1. For example when ACSL1 is overexpressed in rat primary hepatocytes FAs are incorporated into DAG and phospholipids but not into CEorsecretedTAG[11].OverexpressedACSL5increases FA incorporation into DAG CH5424802 and TAG but does not affect FA used for β-oxidation [16]. In contrast however over-expressing either human ACSL3 or rat ACSL5 in HepG2 cells increases palmitate oxidation [17]. Further TAG content increases in NIH-3T3 fibroblasts CH5424802 CH5424802 when ACSL1/ FATP1 is overexpressed [18] and in insulin-stimulated 3T3-L1 adipocytes when ACSL1 is knocked down [13??]. We see two potential problems. The first is that over-expression of an enzyme may overwhelm downstream pathways that cannot readily accommodate excess substrate; the second problem is that studies in cell lines are not always translatable to primary cells. Thus proteomic analysis of mitochondria isolated from white adipose tissue brown adipose CH5424802 tissue and 3T3-L1 adipocytes reveals substantial differences [19??]. Compared with mitochondria from adipose tissue many proteins are downregulated in 3T3-L1 mitochondria. Further medium-chain ACS (ACSM) isoforms are more abundant than ACSL isoforms in white adipose tissue mitochondria suggesting greater medium-chain FA oxidative metabolism. ACSL1 and ACSL6 are particularly enriched in white adipose mitochondria whereas ACSL5 is more abundant in mitochondria from brown adipose. However despite the high abundance of ACSL5 in brown adipose mitochondria ACSL1 is required for adaptive thermo-genesis and normal FA oxidation in brown adipose (J.M. Ellis R.A. Coleman unpublished data). Thus the function of ACSL5 in BAT remains unknown. FA channeling could CH5424802 occur via protein-protein interactions. Immunoprecipitation of endogenous FATP1 from 3T3-L1 adipocytes followed by mass spectrometry identified mitochondrial 2-oxoglutarate dehydrogenase (OGDH) a key enzyme in the tricarboxylic acid cycle [20??]. FATP1 enhances OGDH activity in proteoliposomes whereas FATP1 knockdown in 3T3-L1 adipocytes showed decreased OGDH and TCA cycle.