Figure 3. MPK3/6‐mediated ARR1/10/12 phosphorylation in vitro .

• A–C
  Thiophosphorylation assays of wild‐type and mutated forms ARR1/10/12 by MPK3/6 in vitro. Potential phosphorylated Ser residues or Thr residues of ARR1, ARR10, and ARR12 were mutated to Ala (ARR1^m/ARR10^m/ARR12^m). Recombinant ARR1/10/12‐His and ARR1^m/10^m/12^m‐His proteins were incubated with MPK3 and MPK6 which were activated by MKK5^DD. Phosphorylated ARR1/10/12, ARR1^m/10^m/12^m, and MPK3/6 were visualized by an anti‐thiophosphate ester‐specific antibody (α‐TPE) (upper panel). Asterisk in (B) indicates non‐specific bands. Recombinant MKK5^DD, MPK3, MPK6, ARR1/10/12, and ARR1^m/10^m/12^m were detected by coomassie brilliant blue (CBB) staining as loading controls (bottom panel).

Data information: Two independent biological repeats showed similar results.

Source data are available online for this figure.