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R. E. Carpio, S. Schimpf, E. Zrenner, B. Wissinger; Identification of the CNGA3 Gene Promoter. Invest. Ophthalmol. Vis. Sci. 2008;49(13):1700.
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The cone photoreceptor cyclic nucleotide-gated (CNG) channel is a heterotetramer composed of two alpha and two beta subunits. This channel plays a fundamental role in phototransduction in converting sensory input into electrical responses. Achromatopsia is an inherited autosomal recessive retinal disorder characterized by a lack of cone function and frequently caused by mutations in the CNGA3 gene. Examination of the functional promoter of the CNGA3 gene may provide insights into the mechanisms of cone specific gene expression and reveal important sequences that should be included in mutation screening studies.
5’ RACE experiments were performed with human retinal mRNA. Luciferase reporter gene constructs containing the putative human CNGA3 promoter were transfected into HEK 293, retinoblastoma Y79, WERI-RB-1 cells and 661W cells and promoter activity assayed by luminometry. In addition, interspecies sequence comparison and predictions of putative transcription factor binding sites were done to identify potential proximal promoter sequences.
5’ RACE analysis confirmed the presence of a new first exon of the CNGA3 gene in human retinal tissue that is conserved among mammalian species. Reporter gene assays identified a minimal sequence with promoter function located upstream of exon 1 that yielded significant higher promoter activity compared with other predicted CNGA3 promoters. The highest CNGA3 expression was observed upon transfection of the cone photoreceptor derived 661W mouse cell line.
The results presented here locate the basic CNGA3 gene promoter immediately upstream of exon1 with most significant activity in the cone photoreceptor, derived, mouse 661W cell line.
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