Figure 6

Disruption of the 5' half-site leads to loss of Coup-TFII binding. (A) Electrophoretic mobility shift assay with element C and mutated derivatives (compare (D)). Bands were quantified, background corrected and analyzed. Probe = labelled oligonucleotide used; comp. = unlabelled competitor oligonucleotide used; CII = nuclear extract of Coup-TFII transfected HEK293 cells; mock = nuclear extract of mock transfected HEK293 cells. (B) Quantification of complex intensity on labelled probes Cwt, Cmut5', Cmut3' and Cmut5'3'. Probe Cmut3' retained a lowered ability to bind Coup-TFII while Cmut5'and Cmut5'3'were unable to do so. (C) Accordingly unlabelled competitor probe Cmut3' was still able to partially compete for Coup-TFII binding to the labelled wt probe, while Cmut5' and Cmut5'3' were not. (D) Overview of the probes used in electrophoretic mobility shift assays. Probe Cwt represents element C, the mutated derivatives are depicted below.