Figure 7

Electrophoretic mobility shift assay of RIN binding to the regulatory regions of target genes. (a) SDS-PAGE gel shows the affinity purification of the recombinant RIN protein used for the electrophoretic mobility shift assay. (b) RIN binds specifically to the promoters of target genes containing CArG box elements. The promoter structures of the target genes are presented. Blue boxes indicate CArG box elements in the promoter region and numbers represent the position of these motifs relative to the translational start site. The probe sequences for each target gene are shown, with red letters representing the CArG box. The mutated bases in the probes are represented by blue letters. wt, probe with intact CArG box element; mt, probe with mutated CArG box element. As competitors, 1,000-fold excess amounts of unlabeled probes were added to the binding reaction. The retarded bands and the free probes are indicated by arrowheads.