SC COBRE: REGULATION OF MAMMALIAN FATTY ACID ALPHA-HYDROXYLASE GENE

点击翻译按钮获取中文摘要

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. In higher vertebrates, nerve conduction is greatly facilitated by myelin, a lipid-rich membrane that wraps around the axons. A number of devastating neurodegenerative diseases are known to cause pathological demyelination. Myelin is highly enriched with two galactose-containing sphingolipids, galactosylceramide (GalCer) and sulfatide. A unique feature of these myelin glycosphingolipids is that approximately one half of the fatty acids in GalCer and sulfatide are 2-hydroxy fatty acids. There are no other mammalian tissues that contain such high concentrations of 2-hydroxy fatty acids, suggesting that the 2-hydroxyl group has a unique role in myelin. Although much is known about the role of GalCer and sulfatide in myelination, specific functions of the 2-hydroxyl group in these glycosphingolipids are not known. We hypothesize that 2-hydroxylation of myelin glycosphingolipids is necessary for myelin function. To test our hypothesis, the human fatty acid 2-hydroxylase gene, FA2H, has been cloned, and a highly sensitive GC/MS-based assay system has been developed. These tools will be applied to pursue the following specific aims: 1) to prove that the human FA2H gene product is a fatty acid 2-hydroxylase; 2) to demonstrate that FA2H is highly expressed in brain; 3) to construct a targeting vector to generate FA2H-knockout mice. We have demonstrated increased 2-hydroxy fatty acids and fatty 2-hydroxylase activities in tissue culture cells overexpressing human FA2H. Northern blot analysis showed that FA2H is highly expressed in brain. Mouse brain 2-hydroxy fatty acids and fatty acid 2-hdyroyxlase activities sharply increased during active myelination and decreased as animals aged. These results provide evidence that FA2H-dependent fatty acid 2-hydroxylation is required for the biosynthesis of 2-hydroxy sphingolipids in myelin. Targeting vectors to generate FA2H-knockout mice are currently under construction. With FA2H-knockout mice, we will determine the role of fatty acid 2-hydroxylase in myelination in vivo