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M. E. Morrissey, B. N. Kennedy; CPRE-2 is Sufficient to Enhance Expression From a Cone Photoreceptor-Specific Promoter. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1683.
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© ARVO (1962-2015); The Authors (2016-present)
The alpha subunit of cone transducin (TαC) mediates phototransduction in cone photoreceptors. Mutations in the gene encoding TαC are associated with cone-based blindness. Previously, we demonstrated that ~3.2, ~2.5 and ~0.7 kb promoter fragments of zebrafish TαC are sufficient to drive strong, moderate and weak expression in cone photoreceptors, respectively. The aim of this study is to identify and characterize cis-regulators of cone photoreceptor gene expression.
Using electrophoretic mobility shift assays and transient transgenesis assays, we identified and characterised a regulatory sequence in the proximal promoter of zebrafish TαC.
A 41 bp sequence, designated CPRE-2, located ~0.5 kb upstream of the TαC translation start site was identified by its ability to specifically bind nuclear factors expressed in ocular tissue in vitro. Mutation and deletion analysis shows that the majority of the 41 bp sequence is required for binding of nuclear factors in vitro. In vivo, deletion of CPRE-2 has no effect on the activity levels from a ~2.5 kb TαC promoter fragment. Bioinformatic alignments reveal several CPRE-2 like sequences in the ~2.5 kb promoter suggestive of redundant regulatory sequences. However, three copies of CPRE-2 are sufficient to enhance reporter expression from a minimal UV opsin promoter. Bioinformatic alignments also reveal a CPRE-2 like sequence in the rhodopsin proximal promoter, the function of which is under investigation.
CPRE-2 binds nuclear factors expressed in ocular tissue and it is sufficient to enhance cone photoreceptor gene expression.
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