Hexacosenoyl-CoA is the most abundant very long-chain acyl-CoA in ATP binding cassette transporter D1-deficient cells

Abstract

X-linked adrenoleukodystrophy (X-ALD) is an inherited disorder caused by deleterious mutations in the ABCD1 gene. The ABCD1 protein transports very long-chain FAs (VLCFAs) from the cytosol into the peroxisome where the VLCFAs are degraded through β-oxidation. ABCD1 dysfunction leads to VLCFA accumulation in individuals with X-ALD. FAs are activated by esterification to CoA before metabolic utilization. However, the intracellular pools and metabolic profiles of individual acyl-CoA esters have not been fully analyzed. In this study, we profiled the acyl-CoA species in fibroblasts from X-ALD patients and in ABCD1-deficient HeLa cells. We found that hexacosenoyl (26:1)-CoA, but not hexacosanoyl (26:0)-CoA, was the most abundantly concentrated among the VLCFA-CoA species in these cells. We also show that 26:1-CoA is mainly synthesized from oleoyl-CoA, and the metabolic turnover rate of 26:1-CoA was almost identical to that of oleoyl-CoA in both WT and ABCD1-deficient HeLa cells. The findings of our study provide precise quantitative and metabolic information of each acyl-CoA species in living cells. Our results suggest that VLCFA is endogenously synthesized as VLCFA-CoA through a FA elongation pathway and is then efficiently converted to other metabolites, such as phospholipids, in the absence of ABCD1.

Very long-chain fatty acids are accumulated in triacylglycerol and nonesterified forms in colorectal cancer tissues

“…Analysis was conducted in the positive ion mode using the parallel reaction monitoring (PRM) method. The following conditions were used for positive ion PRM: resolution, 35,000; stepped nce, 30; AGC target, 2e5 ; Maximum IT, 100 ms; Isolation window, 1.0 m/z; ion spray voltage, 1.5 kV; capillary temperature, 250 °C; S-lens RF level 50 V. Nitrogen was used as sheath, auxiliary and collision…”

Non-esterified Very Long-chain Fatty Acids Are Accumulated in Colorectal Cancer Tissues

“…To profile the FFA species as non-esterified fatty acids in CRC tissues, we analyzed the underivatized FFA in total lipid fractions by LC-MS method. We used a C8 reversed-phase column and mobile phases adjusted to pH 9.0 by ammonium hydroxide, which are almost identical with the methods for acyl-CoA analyses developed so far [5]. We validated our method using two isotopically-labeled palmitic acids, and confirmed that 8-2000 pmol/injection of FFA species can be quantified with the present quantitative method ( Supplementary Table S2 online).…”

“…suggesting that FATP4 expression could be different between different types of cancer. Further analysis should be undertaken to clarify So far, we inhibited FATP4 expression by siRNA in ABCD1-deficient HeLa cells, in which PL species and acyl-CoA species harboring VLCFA moiety are accumulated [5]. However, significant difference could not be observed in the amount of non-esterified VLCFA, as compared with control-siRNA treated cells (data not shown).…”

“…2 and Supplementary figure S1, online). It has been shown that the VLCFA moiety is located at the sn-1 position of the glycerol backbone of PL [5,[16][17][18]; thus it is also possible that phospholipase A1 contributes to the accumulation of non-esterified VLCFA in CRC tissues. Three intracellular phospholipase A1 identified so far in mammals (PA-PLA1a/DDHD1/iPLA1a, p125/Se23IP/iPLA1b and KIAA0725p/DDHD2/iPLA1g) prefer PA, PE and PI but not PC as substrates [19].…”

“…Samples were stored at −20°C until further analysis. For acyl-CoA analysis, the tissue specimen were homogenized in 0.9 mL of acetonitrile/isopropanol (3:1 by volume) and the acyl-CoA fraction was extracted as previously described [5]. Samples were stored at −20°C until further analysis.…”