Figure 6

DNAse I footprint analysis of KLF15 binding sites in the rhodopsin promoter. (A) A PCR fragment spanning -225 to +70 bp of the bovine RPPR was end labeled with ³³P at the -225 bp end (forward strand) and incubated with 250, 125, 62.5 and 31.25 ng of purified KLF15-ZF fusion protein (lanes 2–5). Lanes 1 and 6 contained no fusion protein. Protected regions are designated by lines on the left of the image with the nucleotide position of each protected region indicated. Previously identified protein binding sites are indicated by brackets. Solid arrowheads indicate novel hypersensitive sites resulting from KLF15-ZF binding; small arrows indicate hypersensitive sites that are lost or altered by KLF15-ZF binding. Transcription start site (+1) is indicated by an open arrowhead. (B) Same as (A) except the -225 to +70 fragment was labeled at the +70 end (reverse strand). (C) Same as (A) except the PCR fragment used spanned -315 to -31 and was end-labeled with ³³P at the -315 end (forward strand) and lanes 1 and 5 contained no protein. (D) Same as (C) except the -315 to -31 fragment was labeled at the -31 end (reverse strand).