OsSAE1 confers transcriptional repression activity and DNA-binding activity to the GCC-box element. A, Intact and mutated amino acid sequence of the OsSAE1 EAR motif. B, Transient dual-LUC expression assays illustrating the transcriptional repression activity of OsSAE1. The reporters LUC and REN and the effectors (pBD, pBD-OsSAE1, pBD-OsSAE1-EARm, pBD-VP16, and pBD-OsSAE1-VP16) were separately constructed as shown on the left. The activities of LUC and REN were determined 16 h after transformation, and the relative LUC/REN ratios represent the transcriptional activation activities. Error bars represent STDEVs (sds) among three independent replicates. Asterisks indicate significant differences between the compared two samples at *P < 0.05 and **P < 0.01 using a Student’s t test. C, Schematic diagrams of wild-type and deletion mutants of OsSAE1 cloned into the pGEX-6p-1 vector. D, EMSA using GCC-box probe with GST-tagged OsSAE1 (GST-OsSAE1-F) or mutant OsSAE1 (GST-OsSAE1-m1/m2/m3). GST-tag was used in place of GST-OsSAE1 for no-protein controls. Three biological replicates were performed, with similar results.