ELOVL4 Localization to the Endoplasmic Reticulum Is Altered by the Protein Truncation Causing Autosomal Dominant Stargardt-like Macular Dystrophy (STGD3)

Abstract

Abstract: : Purpose: The 5-bp deletion causing STGD3 results in loss of a di-lysine endoplasmic reticulum (ER) retention motif. We hypothesized wild type ELOVL4 would locate to the ER while the mutated STGD3 would be transported out of this compartment. Methods: The ELOVL4 and STGD3 cDNA were expressed by transfection in COS-7 cells as fusion proteins engineered to the carboxyl terminus of enhanced green fluorescence protein (GFP). Transfected cells were stained with fluorescent labeled concanavalin A (ER), wheat germ agglutinin (Golgi apparatus) and Mitotracker Red® CMXRos (mitochondria). GFP-ELOVL4 and GFP-STGD3 subcellular distribution and co-localization with organelle specific stains were assessed by confocal microscopy. Results: GFP-ELOVL4 was expressed as a 64-kDa protein co-localizing with concanavalin A in the endoplasmic reticulum, but not in the Golgi apparatus or mitochondria. The STGD3 mutation resulted in a 56-kDa truncated ELOVL4 protein that failed to localize to the ER, Golgi apparatus, or mitochondria. Surface protease sensitivity demonstrated at least a portion of the mutated STGD3 protein trafficked to the cell surface. Conclusions: The di-lysine retention motif appears to direct ELOVL4 to the endoplasmic reticulum. The STGD3 mutation resulted in failure of the mutant ELOVL4 protein to localize to the ER. Expression of the mutant protein on the cell surface could lead to abnormal photoreceptor physiology and ultimately the RPE dysfunction observed clinically.

Keywords: retinal degenerations: cell biology • retinal degenerations: hereditary • proteins encoded by disease genes