Figure 6.

Systemin and OEs activate LeMPK1 and LeMPK2 in immunocomplex kinase assays. L. peruvianum cells were left untreated (control), or were supplied with systemin or OEs for 10 min (elicitor). Systemin and OE concentrations were as indicated in Figure 1, respectively. Extracts were prepared, incubated with anti-LeMPK1, 2, or 3 antisera (MPK1, 2, 3), and protein G Sepharose. After washing, immunocomplexes were incubated with [γ-³²P]ATP and MBP. Phosphorylated radioactive MBP was separated by SDS-PAGE and was visualized by phosphoimaging. To demonstrate specificity of the anti-LeMPK antisera, antisera were preincubated with an excess of competitor peptides derived from the N-termini of LeMPKs 1, 2, and 3 (peptide 1, 2, and 3) before the addition of extracts.