Figure 5. OsMPK7 interacts and phosphorylates downstream OsWRKY30.

(a) Bimolecular fluorescence assay (BiFc) performed using OsMPK7-eYFP_N173 and OsWRKY30-eYFP_C155 fusion protein reveals interaction between OsMPK7 and OsWRKY30. YFP fluorescence was observed in N. benthamiana cells expressing both OsWRKY30 and OsMPK7 fusion proteins as seen in (a) Epifluorescence, (b) bright field and (c) merged images. No YFP fluorescence is observed in negative controls (d,g and j). Scale bar: 50 μm. (b) In vitro kinase assay showing phosphorylation of OsWRKY30 by its upstream interacting kinase, OsMPK7. OsMPK7 phosphorylated OsWRKY30 in presence of plant expressed OsMKK3-HA protein however bacterially expressed OsMPK7 alone did not phosphorylate OsWRKY30. “+*” Indicate double concentration of OsMPK7. Phosphorylation signal of OsMPK7 and MBP was detected by autoradiography.