PURPOSE: HRG4 (UNC119) is a photoreceptor synaptic protein, a truncation mutant of which has been shown to cause late-onset cone-rod dystrophy in a patient and retinal degeneration with marked synaptic degeneration in a transgenic model. To investigate the mechanism of the retinal degeneration, the effect of the mutant protein expression on the other synaptic proteins was examined. METHODS: The status of 12 known synaptic proteins in the retinas of 5-month- and 13-month-old HRG4 transgenic and control mice was examined by Western blot analysis. Three selected proteins were analyzed by immunofluorescence in the 13-month-old retinas. The 12 proteins were tested for binding to HRG4 by a direct-binding assay and Western blot analysis. RESULTS: A decrease in three synaptic vesicle proteins and an increase in five cytoplasmic and plasma membrane proteins was detected by Western blot analysis in the older but not the younger transgenic retinas. These changes were demonstrated in both the outer and inner plexiform layers of the retina by immunofluorescence, along with a significant reduction in the thickness of the inner plexiform layer. A 23-kDa specie was found to bind to HRG4, but none of the 12 synaptic proteins matched it, according to immunoblot analysis. CONCLUSIONS: The expression of a mutant HRG4 protein in the photoreceptor synapses of the transgenic model had an intrasynaptic and transsynaptic effect, resulting in a decrease in three synaptic vesicle proteins, an increase in five cytoplasmic and plasma membrane proteins, and a significant reduction in the thickness of the inner plexiform layer. These changes were age dependent, similar to the pathologic phenotype of the transgenic model and the patient, and supported a close relationship of HRG4 with other participants in synaptic vesicle function. This interaction was not mediated by a direct coupling of HRG4 with any of the tested synaptic proteins but possibly through interaction with a 23-kDa protein.
PURPOSE:HRG4 (UNC119) is a photoreceptor synaptic protein, a truncation mutant of which has been shown to cause late-onset cone-rod dystrophy in a patient and retinal degeneration with marked synaptic degeneration in a transgenic model. To investigate the mechanism of the retinal degeneration, the effect of the mutant protein expression on the other synaptic proteins was examined. METHODS: The status of 12 known synaptic proteins in the retinas of 5-month- and 13-month-old HRG4 transgenic and control mice was examined by Western blot analysis. Three selected proteins were analyzed by immunofluorescence in the 13-month-old retinas. The 12 proteins were tested for binding to HRG4 by a direct-binding assay and Western blot analysis. RESULTS: A decrease in three synaptic vesicle proteins and an increase in five cytoplasmic and plasma membrane proteins was detected by Western blot analysis in the older but not the younger transgenic retinas. These changes were demonstrated in both the outer and inner plexiform layers of the retina by immunofluorescence, along with a significant reduction in the thickness of the inner plexiform layer. A 23-kDa specie was found to bind to HRG4, but none of the 12 synaptic proteins matched it, according to immunoblot analysis. CONCLUSIONS: The expression of a mutant HRG4 protein in the photoreceptor synapses of the transgenic model had an intrasynaptic and transsynaptic effect, resulting in a decrease in three synaptic vesicle proteins, an increase in five cytoplasmic and plasma membrane proteins, and a significant reduction in the thickness of the inner plexiform layer. These changes were age dependent, similar to the pathologic phenotype of the transgenic model and the patient, and supported a close relationship of HRG4 with other participants in synaptic vesicle function. This interaction was not mediated by a direct coupling of HRG4 with any of the tested synaptic proteins but possibly through interaction with a 23-kDa protein.