Chicco 2018 MiP2018b
Cardiolipin (CL) is a mitochondrial phospholipid that provides structural and functional support to several inner membrane proteins and processes, including the formation of respiratory enzyme supercomplexes. In the healthy human heart, CL is highly enriched with linoleoyl (C18:n2) acyl chains by a remodeling cycle catalyzed by the phospholipid transacylase, tafazzin. Loss-of-function mutations in the tafazzin gene results in linoleoyl-CL deficiency, disrupted supercomplex formation, and cardioskeletal myopathy in Barth syndrome (BTHS). However, it remains unclear exactly how tafazzin and linoleoyl-CL deficiency impact mammalian mitochondrial function and lead to BTHS.
We investigated this using the tafazzin-deficient Taz-shRNA “knockdown” (Taz) mouse, which exhibits deficiencies in linoleoyl-CL and CI-III-IV supercomplexes consistent with BTHS in humans, but have relatively normal cardiac function into early adulthood (suggesting some compensatory adapation). High-resolution respirometry revealed 40-60% lower palmitoylcarnitine- and pyruvate-linked OXPHOS capacity in mitochondria and permeabilized cardiac fibers from Taz vs. wild-type (WT) mice, with only 12-20% lower enzymatic capacities of complexes I, II-IV and IV, and no difference in OXPHOS coupling efficiency or membrane potential. Unexpectedly, glutamate-linked OXPHOS capacity was 50% higher in Taz, and approximated rates of maximal pyruvate and palmitoylcarnitine OXPHOS in WT, arguing against OXPHOS-limiting impairment of the electron transfer system in Taz mitochondria. However, H2O2 emission was elevated during maximal (CI+II) OXPHOS-linked respiration in Taz vs. WT mitochondria, as were oxidation of membrane lipids (4-HNE adducts) and proteins (protein carbonyls).
To investigate the mechanisms of this substrate-specific OXPHOS impairment in Taz mitochondria, cardiac metabolite and mitochondrial proteomic analyses were performed. These studies revealed an accumulation of free amino acids and upregulation of malate dehydrogenase in Taz vs. WT hearts, consistent with an increased reliance on glutamate oxidation to meet energy demands via the malate-aspartate shuttle. Selective deficiencies in free coenzyme-A (CoA), long-chain (>C13) acyl-CoAs, and acetyl-CoA (C2) were also observed, which paralleled changes in long- and medium-chain fatty acid oxidation enzymes and higher acyl-CoA thioesterases. Incubating isolated mitochondria with exogenous CoA restored CoA deficiency and partially rescued pyruvate- and palmitoylcarnitine-supported OXPHOS in Taz, but did not reduce mitochondrial H2O2 emission.
Taken together, these studies suggest that reduced mitochondrial CoA bioavailability, rather than respiratory chain dysfunction, limits carbohydrate and fatty acid OXPHOS in tafazzin-deficient cardiac mitochondria, forcing reliance upon amino acids as an alternative carbon source to meet energy demands. More broadly, the pathogenic impact of impaired mitochondrial supercomplex formation that results from linoleoyl-CL deficiency appears to be increased ROS production, rather than global impairment of OXPHOS capacity or efficiency.
Labels: MiParea: Respiration, nDNA;cell genetics
Organism: Mouse Tissue;cell: Heart Preparation: Permeabilized tissue Enzyme: Complex I, Complex II;succinate dehydrogenase, Complex III, Complex IV;cytochrome c oxidase
Coupling state: OXPHOS Pathway: F, N, NS HRR: Oxygraph-2k
Chicco AJ(1,2), Le CH(2), Benage LG(3), Specht KS(1), Li Puma LC(1), Mulligan CM(1), Heuberger AL(3), Prenni JE(3), Sparagna GC(4)
- Dept Biomedical Sciences
- Cell Molecular Biology Program
- Biochemistry Molecular Biology; Colorado State Univ, Fort Collins
- Division Cardiology, Univ Colorado Denver Anschutz Medical Campus, Aurora; CO, USA. - firstname.lastname@example.org