Fig. 6: Elucidation of VLC-PUFA PC metabolic pathway.

a HeLa cell lipid profiling with VLC-PUFA (FA n-3-32:6) supplementation (n = 4 biologically independent samples). The peak heights of PC 18:1_32:6, DG 18:1_32:6, LPA 32:6, and TG 18:1_18:1_32:6 at final concentrations of 1, 10, or 40 µM of FA n-3-32:6 supplementation are depicted. While LPA was analyzed by a derivatization method using trimethylsilyl-diazomethane, which converts LPA to bis-methyl LPA (BisMeLPA), other molecules were analyzed using conventional untargeted lipidomics methods. Acyl CoA (b) and LPA (c) profiling for the glycerol 3-phosphate acyltransferase 1 (GPAT1) recombinant enzyme assay (n = 4 independently prepared samples). The acyl CoAs and LPAs were analyzed with the vehicle, mock (native plasmid vector), active GPAT1^WT, and the inactive GPAT1 mutant (GPAT1^H230A), supplied with glycerol 3-phosphate and coenzyme in addition to ¹³C-uniformly labeled palmitic acid (FA 16:0 U-¹³C), docosahexaenoic acid (DHA, FA 22:6), or FA n-3-32:6, in the cell-free system enzymatic reaction. The fatty acid was supplied at a final concentration of 10 μM, and the same amount of 17:1 CoA (Fig. 6b) and LPA 17:1 (Fig. 6c) was supplied as the internal standards. The putative ratio between the converted product and the internal standard was used for the y-axis value of dot plots. The mean values were also described in dot plots. Significances were adjusted by false discovery rate in the student’s t-test (two-sided). Source data are provided as a Source Data file.