Figure 4.

Promotion of Cell Death by Phospho-Mimicking Mutations on the SP Cluster of WRKY Transcription Factors.

(A) Phospho-mimicking mutations in a SP cluster. Putative phosphorylated Ser residues in SP cluster were substituted with Asp (nD).

(B) Induction of cell death by overexpression of phospho-mimicking mutants. Leaves were infiltrated with Agrobacterium carrying the indicated gene expression constructs. Photographs were taken 4 d after agroinfiltration (left). After taking the photographs, WRKY-induced cell death was detected by trypan blue staining (right).

(C) Detection of WRKY-HA-StrepII proteins by anti-HA antibody. Total proteins were prepared 48 h after agroinfiltration. Protein loads were monitored by Coomassie blue (CBB) staining of the bands corresponding to the ribulose-1,5-bisphosphate carboxylase large subunit (RBCL). Red asterisks indicate WRKY-HA-StrepII proteins. C, control plant; W, wild-type WRKYs; D, phospho-mimicking mutants.

(D) Detection of WRKY10-HA-StrepII protein. Total proteins were purified by Strep-Tactin and then immunoblot analysis was done.