Mouse very long-chain Acyl-CoA synthetase 3/fatty acid transport protein 3 catalyzes fatty acid activation but not fatty acid transport in MA-10 cells.
The family of proteins that includes very long-chain acyl-CoA synthetases (ACSVL) consists of six members. These enzymes have also been designated fatty acid transport proteins. We cloned full-length mouse Acsvl3 cDNA and characterized its protein product ACSVL3/fatty acid transport protein 3. The predicted amino acid sequence contains two highly conserved motifs characteristic of acyl-CoA synthetases. Northern blot analysis revealed that the mouse Acsvl3 mRNA is highly expressed in adrenal gland, testis, and ovary, with lower expression in the brain of adult mice. A developmental Northern blot revealed that Acsvl3 mRNA levels were significantly higher in embryonic mouse brain (embryonic days 12-14) than in newborn or adult mice, suggesting a possible role in nervous system development. Immunohistochemistry revealed high ACSVL3 expression in adrenal cortical cells, spermatocytes and interstitial cells of the testis, theca cells of the ovary, cerebral cortical neurons, and cerebellar Purkinje cells. Endogenous ACSVL3 was found primarily in mitochondria of MA-10 and Neuro2a cells by both Western blot analysis of subcellular fractions and immunofluorescence analysis. In MA-10 cells, loss-of-function studies using RNA interference confirmed that endogenous ACSVL3 is an acyl-CoA synthetase capable of activating both long-chain (C16:0) and very long-chain (C24:0) fatty acids. However, despite decreased acyl-CoA synthetase activity, initial rates of fatty acid uptake were unaffected by knockdown of Acsvl3 expression in MA-10 cells. These studies cast doubt on the designation of ACSVL3 as a fatty acid transport protein.
Kennedy Krieger Research Institute and Department of Neurology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Zhengtong
Z
Pei
Peter
P
Fraisl
Johannes
J
Berger
Zhenzhen
Z
Jia
Sonja
S
Forss-Petter
Paul A
PA
Watkins
D000311:Adrenal Glands / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D015152:Blotting, Northern; D001921:Brain / Q000196:embryology; D001923:Brain Chemistry; D019556:COS Cells; D002460:Cell Line; D003001:Cloning, Molecular; D003066:Coenzyme A Ligases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism*; D005260:Female; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D005982:Glutathione Transferase / Q000235:genetics; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D009928:Organ Specificity; D010053:Ovary / Q000737:chemistry; D012333:RNA, Messenger / Q000032:analysis; D034741:RNA, Small Interfering / Q000235:genetics; D011993:Recombinant Fusion Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013347:Subcellular Fractions / Q000737:chemistry; D013737:Testis / Q000737:chemistry; D014162:Transfection; D000098859:Long-Chain-Fatty-Acid-CoA Ligase
D005227:Fatty Acids; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D003066:Coenzyme A Ligases; D000098859:Long-Chain-Fatty-Acid-CoA Ligase