Extended Data Fig. 8: The interaction between GST-PAR2 and MBP-MYB27 and the disruption of MYB27–FIE interaction by PAR2.
a, In vitro pull-down assays between GST-PAR2 and MBP-MYB27. The pull-down between GST and MBP-MYB27 was used as the negative control. b, CoIP assays between GFP/MYB27-GFP and PAR2-6Myc in Arabidopsis protoplasts. The proteins were precipitated by anti-GFP beads and detected by anti-GFP or anti-Myc antibody. c, Yeast one-hybrid assay between PAR2 and the 2000 bp SE promoter. Transformed yeast cells were grown on media lacking tryptophan and histidine (SD/-Trp/-His). pHisi.1-0 or pDEST22-0 refers to empty only. 2 mM 3AT was used for self-activation repression. Yeast one-hybrid assay between MYB27 and SE promoter was used as the positive control. d, The comparison of dual-luciferase assays between pSE:LUC plus p35S::GFP and pSE:LUC plus p35S:PAR2 in N. benthamiana leaves (upper panel). The relative intensity of luciferase was quantitively presented by qRT-PCR (lower panel). Values are the means ± s.d. of three biological replicates. e, The binding of PAR2-6Myc at SE promoter (P2) in rosette leaves of par2-1 pPAR2:PAR2-6Myc and par2-1 myb27-1 pPAR2:PAR2-6Myc plants. The y axis shows the calibrated relative ratio of bound DNAs to input DNAs after IP, and IgG serves as the negative control. #1 and #2 represent different individual plants in the same lines. Values are the means ± s.d. of three biological replicates. f, The disruption of MYB27–FIE interaction by PAR2 through yeast-three-hybrid assays. g, The disruption of MYB27–FIE interaction by PAR2 through BiLC assays. In d and e: **p < 0.01; ns: no significance, two-sided Student’s t-test. The exact p values are shown in Supplementary Data 4. The experiments in a and b were repeated independently at least twice with similar results.
