Abstract
Very long chain polyunsaturated fatty acid (VLC-PUFA)-containing glycerophospholipids are highly enriched in the retina; however, details regarding the specific synthesis and function of these highly unusual retinal glycerophospholipids are lacking. Elongation of very long chain fatty acids-4 (ELOVL4) has been identified as a fatty acid elongase protein involved in the synthesis of VLC-PUFAs. Mutations in ELOVL4 have also been implicated in an autosomal dominant form of Stargardt disease (STGD3), a type of juvenile macular degeneration. We have generated photoreceptor-specific conditional knock-out mice and used high performance liquid chromatography-mass spectrometry (HPLC-MS) to examine and analyze the fatty acid composition of retinal membrane glycerophosphatidylcholine and glycerophosphatidylethanolamine species. We also used immunofluorescent staining and histology coupled with electrophysiological data to assess retinal morphology and visual response. The conditional knock-out mice showed a significant decrease in retinal glycerophospholipids containing VLC-PUFAs, specifically contained in the sn-1 position of glycerophosphatidylcholine, implicating the role of Elovl4 in their synthesis. Conditional knock-out mice were also found to have abnormal accumulation of lipid droplets and lipofuscin-like granules while demonstrating photoreceptor-specific abnormalities in visual response, indicating the critical role of Elovl4 for proper rod or cone photoreceptor function. Altogether, this study demonstrates the essential role of ELOVL4 in VLC-PUFA synthesis and retinal function.
Highlights
Phospholipids containing very long chain polyunsaturated fatty acids (VLC-PUFAs) are enriched in retina
Analysis of Retinal Extract Glycerophospholipids—Extracts obtained from wild type (WT) mouse retina were analyzed for a total of 21 different PC and PC and glycerophosphoethanolamine (PE) molecular species (42 total), including those containing both five- and six-double bond VLC-PUFA acyl chains; these results are shown in Fig. 1, C and D
An high performance liquid chromatography-mass spectrometry (HPLC-mass spectrometry (MS)) multiple reaction monitoring (MRM) method was created that included precursor anion mass/acyl chain fragment anion mass MRM pairs specific for these 21 PC and PE species
Summary
Phospholipids containing very long chain polyunsaturated fatty acids (VLC-PUFAs) are enriched in retina. Elongation of very long chain fatty acids-4 (ELOVL4) has been identified as a fatty acid elongase protein involved in the synthesis of VLC-PUFAs. Mutations in ELOVL4 have been implicated in an autosomal dominant form of Stargardt disease (STGD3), a type of juvenile macular degeneration. Cameron et al [22] examined the specific elongase activity of ELOVL4 using homozygous Elovl Y270X knock-in and homozygous Elovl knock-out mice and observed that mice lacking functional ELOVL4 died perinatally due in part to dehydration from a faulty skin permeability barrier The skin of both the homozygous mutant and the knock-out mouse was deficient in glycerophospholipids containing fatty acids 26 carbon atoms or longer in chain length, implicating ELOVL4 in their biosynthesis. Our results indicate that ELOVL4 is essential for VLCPUFA synthesis as well as retinal functions
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