Extended Data Fig. 7: Effect of Trx-h2 on the binding of CBF1 to the COR15a promoter and on the expression of CBF mRNAs and proteins at 4 °C.
From: Redox-dependent structural switch and CBF activation confer freezing tolerance in plants
a, Nucleotide sequence of the COR15a promoter containing two CRT/DRE core motifs at nucleotide positions -439 to -444 (blue box) and -267 to -262 (green box) upstream of the transcription start site (+1 bp). b, Schematic representation of the COR15a promoter containing two CRT/DRE cis-elements indicated as blue and green boxes. c, Oligonucleotide sequence of biotin-labeled EMSA probe (-275 to -253 bp). d, Schematic representation of effector, internal control, and reporter constructs used in the luciferase (LUC) assay. e, Comparison of LUC activity measured in Nicotiana benthamiana leaves expressing P35S:CBF1 under oxidizing (X/XO) or reducing (GSH) conditions at 22 °C. f, Comparison of LUC activity in N. benthamiana leaves expressing P35S:Trx-h2 or P35S:Trx-h2(C/S) incubated at 22 °C or 4 °C. g, Analysis of CBF1–3 transcript levels in Col-0, trx-h2, and trx-h3 (control) plants incubated at 4 °C by qRT-PCR. Data are expressed as mean ± s.e.m (n = 3 biologically independent samples). Significant differences between means are indicated by asterisks (Unpaired two-tailed Student’s t test P values: *P < 0.05, **P < 0.01, ***P < 0.001). NS indicates not significant. h, Expression analysis of CBF proteins by western blot on reducing SDS-PAGE gels in Col-0, trx-h2, and cbfs Arabidopsis during the cold treatment at 4 °C. Rbc L stained with Ponceau S was used as loading controls.
