Figure 7.

In Vivo Phosphorylation of WRKY33 by MPK3/MPK6.

(A) WRKY33 becomes phosphorylated in seedlings infected with B. cinerea. Protein extracts from wild-type (Col-0), wrky33/4myc-WRKY33^WT, and wrky33/4myc-WRKY33^SA seedlings treated with B. cinerea for different times were separated in an SDS-PAGE gel with Phos-tag reagent. After being transferred to a nitrocellulose membrane, myc-tagged WRKY33^WT and WRKY33^SA proteins were detected by an anti-myc antibody (top). A regular immunoblot (IB) was done at the same time to detect total WRKY33 protein (middle). Equal loading was confirmed by Ponceau S staining (bottom).

(B) WRKY33 phosphorylation in gain-of-function DD seedlings after DEX treatment. Protein extracts from DD, DD/wrky33/4myc-WRKY33^WT, and DD/wrky33/4myc-WRKY33^SA seedlings treated with DEX (1 μM) at various times were separated in a Phos-tag SDS-PAGE gel. After being transferred to nitrocellulose membranes, WRKY33^WT and WRKY33^SA proteins were detected by an anti-myc antibody (top). A regular immunoblot was done at the same time to detect the total WRKY33 protein (middle). Equal loading was confirmed by Ponceau S staining (bottom).