Cardiolipin-induced activation of pyruvate dehydrogenase links mitochondrial lipid biosynthesis to TCA cycle function

Abstract

Edited by George M. Carman Cardiolipin (CL) is the signature phospholipid of mitochondrial membranes. Although it has long been known that CL plays an important role in mitochondrial bioenergetics, recent evidence in the yeast model indicates that CL is also essential for intermediary metabolism. To gain insight into the function of CL in energy metabolism in mammalian cells, here we analyzed the metabolic flux of [U-13 C]glucose in a mouse C2C12 myoblast cell line, TAZ-KO, which is CL-deficient because of CRISPR/Cas9-mediated knockout of the CL-remodeling enzyme tafazzin (TAZ). TAZ-KO cells exhibited decreased flux of [U-13 C]glucose to [ 13 C]acetyl-CoA and M2 and M4 isotopomers of tricarboxylic acid (TCA) cycle intermediates. The activity of pyruvate carboxylase, the predominant enzyme for anaplerotic replenishing of the TCA cycle, was elevated in TAZ-KO cells, which also exhibited increased sensitivity to the pyruvate carboxylase inhibitor phenylacetate. We attributed a decreased carbon flux from glucose to acetyl-CoA in the TAZ-KO cells to a ϳ50% decrease in pyruvate dehydrogenase (PDH) activity, which was observed in both TAZ-KO cells and cardiac tissue from TAZ-KO mice. Protein-lipid overlay experiments revealed that PDH binds to CL, and supplementing digitonin-solubilized TAZ-KO mitochondria with CL restored PDH activity to WT levels. Mitochondria from TAZ-KO cells exhibited an increase in phosphorylated PDH, levels of which were reduced in the presence of supplemented CL. These findings indicate that CL is required for optimal PDH activation, generation of acetyl-CoA, and TCA cycle function, findings that link the key mitochondrial lipid CL to TCA cycle function and energy metabolism. The phospholipid cardiolipin (CL) 8 is the signature lipid of mitochondrial membranes. It interacts with a wide variety of mitochondrial proteins (1) and has multiple roles in mitochondrial biogenesis as well as other cellular functions (2-4). Loss of CL leads to deficiencies in respiratory function, mitochondrial membrane potential, and ATP synthesis (1, 5-7). CL that is synthesized de novo has predominantly saturated fatty acids (8). However, the lipid is then remodeled via phospholipase-mediated deacylation to form monolysocardiolipin (MLCL), which is subsequently acylated by the enzyme tafazzin to synthesize CL with predominantly unsaturated fatty acids (9, 10). Abnormal CL composition is associated with several human disorders, including diabetes, Alzheimer's disease, and Parkinson's disease (11-13). However, Barth syndrome (BTHS) is the only disorder identified to date that is caused by altered CL metabolism (14, 15). BTHS is a severe X-linked disorder characterized by dilated cardiomyopathy, skeletal myopathy, neutropenia, exercise intolerance, lactic acidosis, and sudden death from arrhythmia (16-18). The underlying cause of BTHS is mutations in the tafazzin (TAZ) gene, which codes for the transacylase that remodels CL (10, 19). As a result, BTHS patients exhibit an aberrant CL profile characterized by decreased tot...

Tafazzin deficiency impairs CoA-dependent oxidative metabolism in cardiac mitochondria

“…Results from the present study suggest a potential block in medium chain acyl-CoA oxidation in Taz KD mitochondria, given evidence for similar impairment of both long-and medium-chain acylcarnitine oxidation, lower MCAD expression, and a relative accumulation of C3-C12 acyl-CoAs. Protein levels of the pyruvate dehydrogenase complex subunits were not lower in Taz KD , but this enzyme complex has been previously shown to be inhibited by posttranslational modifications in this and other Taz-deficient models (55). However, partial rescue of both palmitoylcarnitine-and pyruvatesupported OXPHOS flux with exogenous CoA herein suggests additional limitations imposed by free CoA availability or impaired CoA-dependent reactions.…”

Barth Syndrome: Exploring Cardiac Metabolism with Induced Pluripotent Stem Cell-Derived Cardiomyocytes

“…Our findings, however, are contradictory to the studies with other cellular models. Li et al report reduced carbon flux from glucose to Krebs cycle intermediates in TAZ-KO mouse C2C12 myoblast cell line [54], while Chatzispyrou et al demonstrate that the fractional contribution of glucose to the Krebs cycle intermediates is unaffected in TAZ-deficient skin fibroblasts [55]. The discrepancy between studies can be explained by differences in genotype and cellular metabolism for different types of cells.…”

Cardiolipin, the Mitochondrial Signature Lipid: Implication in Cancer

“…CL also seems required for acetyl CoA synthesis and is essential for the normal function of the tricarboxylic acid (TCA) cycle [60,61]. In this line, the CL-deficient model of a mouse C2C12 myoblast cell line has shown a decreased carbon flux from glucose to acetyl CoA associated with a decrease in pyruvate dehydrogenase activity [60]. Moreover, the implication of acetyl CoA in cancer cell migration and metastasis has already been demonstrated [62][63][64][65][66].…”