Conditional Ablation of Retinal Elovl4 Reveals a Key Role in Synthesis of VLC-PUFAs and Photoreceptor Light Responses

Abstract

 

Purpose:

 

To compare biochemical, physiological and behavioral phenotypes incurred by the genetic ablation of the Elovl4 elongase and by Elovl4 mutations that impair trafficking of the Elovl4 protein in the transgenic mouse model for autosomal dominant Stargardt disease type 3 (STGD3).

 

Methods:

 

Rod and cone conditional knockout (cKO) mice were generated using the pLox/Cre system. WT1 and TG2 lines carried the human wild type or the human mutant allele (delAACTT at 790-794) in two copies, respectively. RT-PCR and IHC were used to characterize expression and localization of Cre recombinase and Elovl4. Visual acuity and outer retinal physiology were assessed using the optomotor head tracking response and electroretinographic (ERG) analysis, respectively. Gas chromatography coupled with mass spectrometry (GC-MS) was used to determine retinal levels of docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA) and C₂₄-C₃₄ very long chain polyunsaturated fatty acids (VLC-PUFAs).

 

Results:

 

DHA and C₂₄ lipid content of rod cKO retinas did not change whereas there was a ~ 50% decrease in C₃₀-C₃₄ VLC-PUFAs. In contrast, a 43.3% decrease in DHA and 66.5% decrease in C24:5n3 content was measured in TG2 retinas together with undetectable content of C₃₀-C₃₄ VLC-PUFAs. TG2 mice showed a severe visual phenotype with visual acuity that decreased from 0.380±0.006 cyc/deg at 1 month to 0.046±0.030 cyc/deg at 7 months of age. In cone cKO mice, the photopic ERG a- and b-wave decreased by 42.6±11.8% and by 19.2±2.3%, respectively. No ERG changes were observed in rod cKO animals.

 

Conclusions:

 

Our data shows that Elovl4, expressed in rod photoreceptors, is required for synthesis of retinal VLC-PUFAs in the C₃₀-C₃₄ range, but is not required for biosynthesis of C₂₀, C₂₄ lipids. Elovl4 elimination from cones compromised the photopic ERG. The phenotype of transgenic mice carrying the human mutant allele was markedly more severe than the phenotype of mice lacking the Elovl4 protein. We conclude that cone but not rod, retinal signaling is impaired by loss of Elovl4. Moreover, STGD3 may be associated with wider loss of PUFAs and vision loss than expected from a simple loss of Elovl4 protein.

 

Keywords: lipids • retinal degenerations: hereditary • proteins encoded by disease genes