Acetyl‐CoA generated in peroxisomes of CHO and HepG2 cells is preferentially incorporated into sterols versus fatty acids: studies with [U‐13C12]dodecanedioate

Abstract

When labeled mitochondrial acetyl-CoA is exported to the cytosol via citrate, acetyl groups incorporated into fatty acids and sterols are equally labeled. We compared the mass isotopomer distributions (MID) of fatty acids and sterols in CHO and HepG2 cells incubated with 2 mM [U-13C12]dodecanedioate for 48 or 72 hr. The initial steps of the B-oxidation of this dicarboxylate generate highly labeled acetyl-CoA in peroxisomes. As controls, we incubated HepG2 cells with 25 mM [U-13C6]glucose, which forms mitochondrial [U-13C2]acetyl CoA. The MID of fatty acids (palmitate) and sterols (cholesterol) were subjected to Isotopomer Spectral Analysis to estimate the fractional contribution of the labeled precursor to acetyl CoA. The labeling ratio (acetyl of sterols)/(acetyl of fatty acids) was 1.04 + 0.09 SD. (n = 6) for [13C]glucose. In contrast, in the presence of [U-13C12]dodecanedioate, the labeling ratio was much greater than 1, i.e., 1.72 in CHO and 1.30 in HepG2 cells. Fractional synthesis of fatty acids and sterols varied from 23% to 95%. However, the elevated sterol to fatty acid ratio for acetyl CoA enrichment was maintained under all conditions for [U-13C12]dodecanedioate compared to [U-13C6]glucose. Our data are consistent with the hypothesis that part of sterol synthesis in CHO and HepG2 cells occurs in peroxisomes. Supported by NIH Roadmap grant R33DK070291.