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Novel role of FATP1 in mitochondrial fatty acid oxidation in skeletal muscle cells.

Sebastián D, Guitart M, García-Martínez C, Mauvezin C, Orellana-Gavaldà JM, Serra D, Gómez-Foix AM, Hegardt FG, Asins G - J. Lipid Res. (2009)

Bottom Line: The cooverexpression of FATP1 and CPT1 also enhanced mitochondrial fatty acid oxidation, similar to the cooverexpression of FAT/CD36 and CPT1.However, etomoxir, an irreversible inhibitor of CPT1, blocked all these effects.These data reveal that FATP1, like FAT/CD36, is associated with mitochondria and has a role in mitochondrial oxidation of fatty acids.

View Article: PubMed Central - PubMed

Affiliation: Departments of Biochemistry and Molecular Biology, School of Pharmacy, University of Barcelona, Barcelona, Spain.

ABSTRACT
Carnitine palmitoyltransferase 1 (CPT1) catalyzes the first step in long-chain fatty acid import into mitochondria, and it is believed to be rate limiting for beta-oxidation of fatty acids. However, in muscle, other proteins may collaborate with CPT1. Fatty acid translocase/CD36 (FAT/CD36) may interact with CPT1 and contribute to fatty acid import into mitochondria in muscle. Here, we demonstrate that another membrane-bound fatty acid binding protein, fatty acid transport protein 1 (FATP1), collaborates with CPT1 for fatty acid import into mitochondria. Overexpression of FATP1 using adenovirus in L6E9 myotubes increased both fatty acid oxidation and palmitate esterification into triacylglycerides. Moreover, immunocytochemistry assays in transfected L6E9 myotubes showed that FATP1 was present in mitochondria and coimmunoprecipitated with CPT1 in L6E9 myotubes and rat skeletal muscle in vivo. The cooverexpression of FATP1 and CPT1 also enhanced mitochondrial fatty acid oxidation, similar to the cooverexpression of FAT/CD36 and CPT1. However, etomoxir, an irreversible inhibitor of CPT1, blocked all these effects. These data reveal that FATP1, like FAT/CD36, is associated with mitochondria and has a role in mitochondrial oxidation of fatty acids.

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Overexpression of FATP1, FAT/CD36, and CPT1A in L6E9 myotubes. L6E9 myotubes were transduced with Ad-FATP1, Ad-FAT/CD36, Ad-CPT1A, or Ad-LacZ as a control, and a Western blot using a specific antibody against FATP1 (A), FAT/CD36 (B), or CPT1A (C) was performed in 50 μg of total extract.
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fig1: Overexpression of FATP1, FAT/CD36, and CPT1A in L6E9 myotubes. L6E9 myotubes were transduced with Ad-FATP1, Ad-FAT/CD36, Ad-CPT1A, or Ad-LacZ as a control, and a Western blot using a specific antibody against FATP1 (A), FAT/CD36 (B), or CPT1A (C) was performed in 50 μg of total extract.

Mentions: Mouse FATP1 cDNA was delivered into L6E9 myotubes by adenovirus. FATP1 protein levels increased 2-fold in Ad-FATP1-transduced cells compared with control Ad-LacZ-transduced cells (Fig. 1A). To assess the functionality of the overexpressed FATP1, we measured the incorporation of palmitate into lipids and oxidation in intact cells. FATP1 overexpression produced a 2-fold increase in the incorporation of palmitate into TGs (LacZ control value: 0.36 ± 0.04 nmol palmitate/mg protein × h), and no increase was seen for PLs, DAG, or nonesterified palmitate (NEPalm) (LacZ control values in nmol palmitate/mg protein × h: 0.14 ± 0.01, 0.08 ± 0.01, and 0.033 ± 0.003 for PL, DAG, and NEPalm, respectively; Fig. 2A). L6E9 myotubes transduced with Ad-FATP1 showed a 1.5-fold increase in palmitate oxidation to CO2 compared with Ad-LacZ-transduced myotubes (6.5 ± 0.27 nmol palmitate/mg protein × 3 h; Fig. 2B). We also examined the effect of FAT/CD36 overexpression in L6E9 myotubes, a protein that has already been shown to have a role in fatty acid oxidation in muscle (12). Overexpression of FAT/CD36 (2-fold increase in protein levels; Fig. 1B) produced a 1.5- to 1.8-fold increase in palmitate esterification to TG, PL, DAG, and NEPalm (Fig. 2A) and a 1.5-fold increase in palmitate oxidation (Fig. 2B).


Novel role of FATP1 in mitochondrial fatty acid oxidation in skeletal muscle cells.

Sebastián D, Guitart M, García-Martínez C, Mauvezin C, Orellana-Gavaldà JM, Serra D, Gómez-Foix AM, Hegardt FG, Asins G - J. Lipid Res. (2009)

Overexpression of FATP1, FAT/CD36, and CPT1A in L6E9 myotubes. L6E9 myotubes were transduced with Ad-FATP1, Ad-FAT/CD36, Ad-CPT1A, or Ad-LacZ as a control, and a Western blot using a specific antibody against FATP1 (A), FAT/CD36 (B), or CPT1A (C) was performed in 50 μg of total extract.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2724792&req=5

fig1: Overexpression of FATP1, FAT/CD36, and CPT1A in L6E9 myotubes. L6E9 myotubes were transduced with Ad-FATP1, Ad-FAT/CD36, Ad-CPT1A, or Ad-LacZ as a control, and a Western blot using a specific antibody against FATP1 (A), FAT/CD36 (B), or CPT1A (C) was performed in 50 μg of total extract.
Mentions: Mouse FATP1 cDNA was delivered into L6E9 myotubes by adenovirus. FATP1 protein levels increased 2-fold in Ad-FATP1-transduced cells compared with control Ad-LacZ-transduced cells (Fig. 1A). To assess the functionality of the overexpressed FATP1, we measured the incorporation of palmitate into lipids and oxidation in intact cells. FATP1 overexpression produced a 2-fold increase in the incorporation of palmitate into TGs (LacZ control value: 0.36 ± 0.04 nmol palmitate/mg protein × h), and no increase was seen for PLs, DAG, or nonesterified palmitate (NEPalm) (LacZ control values in nmol palmitate/mg protein × h: 0.14 ± 0.01, 0.08 ± 0.01, and 0.033 ± 0.003 for PL, DAG, and NEPalm, respectively; Fig. 2A). L6E9 myotubes transduced with Ad-FATP1 showed a 1.5-fold increase in palmitate oxidation to CO2 compared with Ad-LacZ-transduced myotubes (6.5 ± 0.27 nmol palmitate/mg protein × 3 h; Fig. 2B). We also examined the effect of FAT/CD36 overexpression in L6E9 myotubes, a protein that has already been shown to have a role in fatty acid oxidation in muscle (12). Overexpression of FAT/CD36 (2-fold increase in protein levels; Fig. 1B) produced a 1.5- to 1.8-fold increase in palmitate esterification to TG, PL, DAG, and NEPalm (Fig. 2A) and a 1.5-fold increase in palmitate oxidation (Fig. 2B).

Bottom Line: The cooverexpression of FATP1 and CPT1 also enhanced mitochondrial fatty acid oxidation, similar to the cooverexpression of FAT/CD36 and CPT1.However, etomoxir, an irreversible inhibitor of CPT1, blocked all these effects.These data reveal that FATP1, like FAT/CD36, is associated with mitochondria and has a role in mitochondrial oxidation of fatty acids.

View Article: PubMed Central - PubMed

Affiliation: Departments of Biochemistry and Molecular Biology, School of Pharmacy, University of Barcelona, Barcelona, Spain.

ABSTRACT
Carnitine palmitoyltransferase 1 (CPT1) catalyzes the first step in long-chain fatty acid import into mitochondria, and it is believed to be rate limiting for beta-oxidation of fatty acids. However, in muscle, other proteins may collaborate with CPT1. Fatty acid translocase/CD36 (FAT/CD36) may interact with CPT1 and contribute to fatty acid import into mitochondria in muscle. Here, we demonstrate that another membrane-bound fatty acid binding protein, fatty acid transport protein 1 (FATP1), collaborates with CPT1 for fatty acid import into mitochondria. Overexpression of FATP1 using adenovirus in L6E9 myotubes increased both fatty acid oxidation and palmitate esterification into triacylglycerides. Moreover, immunocytochemistry assays in transfected L6E9 myotubes showed that FATP1 was present in mitochondria and coimmunoprecipitated with CPT1 in L6E9 myotubes and rat skeletal muscle in vivo. The cooverexpression of FATP1 and CPT1 also enhanced mitochondrial fatty acid oxidation, similar to the cooverexpression of FAT/CD36 and CPT1. However, etomoxir, an irreversible inhibitor of CPT1, blocked all these effects. These data reveal that FATP1, like FAT/CD36, is associated with mitochondria and has a role in mitochondrial oxidation of fatty acids.

Show MeSH
Related in: MedlinePlus