PANK2-mediated de novo CoA synthesis is required for metabolic switching to fatty acid oxidation

Pantothenate phosphorylation by pantothenate kinase (PANK) represents the first step in the de novo synthesis of Coenzyme A (CoA). Humans have 4 different PANK enzymes (PANK1-4) although PANK4 was recently shown to act as a phosphatase. Only PANK2 has been detected in the mitochondria. PANKs 1-3 are feedback inhibited by the accumulation of acyl-CoAs. However, PANK2 can overcome this inhibition by binding palmitoyl-carnitine in vitro . Previous studies, conducted under glucose-replete conditions, have failed to detect a PANK2-mediated contribution to CoA synthesis since neither PANK2 deletion nor overexpression leads to changes in CoA cellular content. We found that exposure to BSA-conjugated palmitate (PAL-BSA) led to accumulation of both palmitoyl-carnitine and palmitoyl-CoA without evidence of lipotoxicity in HEK293T cells, suggesting that PANK2 was active under these conditions. Isotope tracing experiments with ¹³ C ¹⁵ N-pantothenate showed a significant drop of de novo CoA [m+4] synthesis in PANK2 silenced (siPANK2) cells that was not observed in control or siPANK1 cells. Newly synthesized CoA in control and siPANK1 cells was able to sustain the high production of acetyl-CoA known to occur with metabolic switching to preferential fatty acid oxidation (FAO). In contrast, PANK2-deficient cells failed to increase acetyl-CoA production during PAL-BSA, indicating that FAO is limited by CoA availability in these conditions. Consistently, we observed diminished cellular viability in PAL-BSA-treated siPANK2 cells concomitant with a drop in ATP levels and the activation of starvation biomarkers AMPK and autophagy. These findings reveal a crucial role for PANK2 in CoA synthesis during fatty acid oxidation.