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. 2023 Feb 11;4(4):100559. doi: 10.1016/j.xplc.2023.100559

Figure 5.

Figure 5

Analyses of the autophosphorylation sites in the activation segment of FER-KD.

(A) Liquid chromatography tandem mass spectrometry identification of FER-KD (green) autophosphorylation sites in vitro at different time points. The activation segment is presented in yellow. The phosphorylation sites are numbered in the top panel. Compared with the previous time point, newly emerging phosphorylation sites are shown in red.

(B) Western blot analysis of autophosphorylation by the FER-KD mutants. Samples (1 mM) were incubated with 1 mM ATP and 10 mM at 25°C for 30 min.

(C) View of the interaction networks of D661, K663, S701, and Y704 in the FER-KD activation segment. The interacting residues are indicated by yellow or green sticks. Hydrogen bonds are presented as black dashed lines.

(D) Identification of the phosphorylation site Y704 in vivo. RALF1-induced phosphorylation of Y704 of FER in 7-day-old pFER::FER-GFP seedlings is shown. Specific bands were recognized by anti-GFP and anti-pY704 antibodies at the indicated periods.