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. 2013 Mar 22;25(3):1126–1142. doi: 10.1105/tpc.112.109074

Figure 1.

Figure 1.

MPK3 and MPK6 Phosphorylate ERF6 on the C-Terminal Motif That Shares Homology with the C Terminus of ACS6.

(A) Alignment of the C termini of ACS6 and ERF6. Plus symbols indicate the conserved MAPK phosphorylation sites.

(B) Diagram of ERF6 protein with the putative MAPK phosphorylation sites marked.

(C) Phosphorylation of ERF6 recombinant protein by activated MPK3 and MPK6. Activated MPK3 and MPK6 were used to phosphorylate purified double-tagged ERF6 (top panel). Side-by-side control reactions using myelin basic protein (MBP) as a substrate validated the activation of MPK3 and MPK6 by the constitutively active MKK4 and MKK5 (bottom panel).

(D) Identification of the MPK3 and MPK6 phosphorylation sites in ERF6 based on in vitro phosphorylation assay. Single and high-order Ser/Thr-to-Ala mutant ERF6 proteins were purified and normalized to the same concentration as validated by Coomassie blue staining (top panel). The ability of ERF6 and its mutants to be phosphorylated by MPK3 (middle panel) and MPK6 (bottom panel) was determined by in vitro phosphorylation assays.