TY - JOUR
T1 - Carnitine palmitoyltransferase 2 and carnitine/acylcarnitine translocase are involved in the mitochondrial synthesis and export of acylcarnitines
AU - Violante, Sara
AU - Ijlst, Lodewijk
AU - te Brinke, Heleen
AU - Tavares de Almeida, Isabel
AU - Wanders, Ronald J. A.
AU - Ventura, Fátima V.
AU - Houten, Sander M.
PY - 2013
Y1 - 2013
N2 - Acylcarnitines are commonly used in the diagnosis of mitochondrial fatty acid β-oxidation disorders (mFAODs). It is generally assumed that this plasma acylcarnitine profile reflects the mitochondrial accumulation of acyl-CoAs. The identity of the enzymes and the mitochondrial and plasmalemmal transporters involved in the synthesis and export of these metabolites have remained undefined. We used lentiviral shRNA to knock down the expression of medium-chain acyl-CoA dehydrogenase (MCAD) in control and carnitine palmitoyltransferase 2 (CPT2)-, carnitine/acylcarnitine translocase (CACT)-, and plasmalemmal carnitine transporter (OCTN2)-deficient human fibroblasts. These cell lines, including mock-transduced controls, were loaded with decanoic acid and carnitine, followed by the measurement of the acylcarnitine profile in the extracellular medium. In control fibroblasts, MCAD knockdown markedly increased the production of octanoylcarnitine (3-fold, P <0.01). OCTN2-deficient cell lines also showed extracellular accumulation of octanoylcarnitine (2.8-fold, P <0.01), suggesting that the cellular export of acylcarnitines does not depend on OCTN2. In contrast, in CPT2- and CACT-deficient cells, the accumulation of octanoylcarnitine in the medium did not significantly increase in the MCAD knockdown. Similar results were obtained using pharmacological inhibition of CPT2 in fibroblasts from MCAD-deficient individuals. This shows that CPT2 and CACT are crucial for mitochondrial acylcarnitine formation and export to the extracellular fluids in mFAOD
AB - Acylcarnitines are commonly used in the diagnosis of mitochondrial fatty acid β-oxidation disorders (mFAODs). It is generally assumed that this plasma acylcarnitine profile reflects the mitochondrial accumulation of acyl-CoAs. The identity of the enzymes and the mitochondrial and plasmalemmal transporters involved in the synthesis and export of these metabolites have remained undefined. We used lentiviral shRNA to knock down the expression of medium-chain acyl-CoA dehydrogenase (MCAD) in control and carnitine palmitoyltransferase 2 (CPT2)-, carnitine/acylcarnitine translocase (CACT)-, and plasmalemmal carnitine transporter (OCTN2)-deficient human fibroblasts. These cell lines, including mock-transduced controls, were loaded with decanoic acid and carnitine, followed by the measurement of the acylcarnitine profile in the extracellular medium. In control fibroblasts, MCAD knockdown markedly increased the production of octanoylcarnitine (3-fold, P <0.01). OCTN2-deficient cell lines also showed extracellular accumulation of octanoylcarnitine (2.8-fold, P <0.01), suggesting that the cellular export of acylcarnitines does not depend on OCTN2. In contrast, in CPT2- and CACT-deficient cells, the accumulation of octanoylcarnitine in the medium did not significantly increase in the MCAD knockdown. Similar results were obtained using pharmacological inhibition of CPT2 in fibroblasts from MCAD-deficient individuals. This shows that CPT2 and CACT are crucial for mitochondrial acylcarnitine formation and export to the extracellular fluids in mFAOD
U2 - https://doi.org/10.1096/fj.12-216689
DO - https://doi.org/10.1096/fj.12-216689
M3 - Article
C2 - 23322164
SN - 0892-6638
VL - 27
SP - 2039
EP - 2044
JO - FASEB Journal
JF - FASEB Journal
IS - 5
ER -