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. 2020 Aug 18;11:1265. doi: 10.3389/fpls.2020.01265

Figure 1.

Figure 1

Identification of ZBED-interacting proteins. (A) Screening of two rice yeast two-hybrid cDNA libraries (CO39 and RCL1) with ZBED led to the identification of several putative interacting partners. WRKY4, STE11, STE20, and KIP1 were found to interact with ZBED in yeast cells. Yeasts co-expressing BD-ZBED with either AD-WRKY4, AD-STE11, AD-STE20, or AD-KIP1 were plated on selective media (SD-LWH) with different concentrations of 3AT. Empty pGBKT7-BD (BD-empty) was used as negative bait control. (B) Bimolecular Fluorescent Complementation (BiFC) assay was used to confirm yeast two-hybrid interactions (right panels). Transient co-expression of YFPc-ZBED either with YFPn-KIP1, YFPn-STE20, and YFPn-WRKY4, led to the detection of a nuclear fluorescent signal. Lack of YFP fluorescence upon co-expression of YFPc-ZBED with YFPn-STE11 suggests that these two proteins do not associate in planta. DAPI staining was used for nuclear staining (left panels). Observations were performed on N. benthamiana leaf samples collected three and four days post infiltration (dpi).