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. Author manuscript; available in PMC: 2016 Feb 26.
Published in final edited form as: Cell Host Microbe. 2014 Oct 8;16(4):473–483. doi: 10.1016/j.chom.2014.09.007

Figure 5. RIN4 Thr166 Phosphorylation Weakens the ROC1-RIN4 Interaction.

Figure 5

(A) The pmRIN4 mutation reduces the ROC1-RIN4 interaction in vitro. His-RIN4 or His-pmRIN4 was incubated with GST, or GST-ROC1 recombinant protein for pull-down assay, and amounts of proteins in the blot were determined by immune blot or CBB staining.

(B) AvrB diminishes the ROC1-RIN4 interaction in Nb plants, whereas AvrAC restores the interaction. The indicated Nluc and Cluc constructs along with AvrB-FLAG, AvrAC-HA, and mAvrAC-HA were transiently expressed in Nb plants for luciferase complementation assay. The experiment was performed twice with similar results. Error bars represent SD (n = 8; 2 biological repreats).

(C) The AvrB-induced RIN4 phosphorylation was diminished in ROC1S58F plants. WT (Col-0) and ROC1S58F plants were infiltrated with Pst containing an empty vector pVSP61 (EV) or avrB, and protein was extracted 2 hr after the onset of HR for immune blot analyses. CBB stain of Rubisco indicate equal loading of total protein. See also Figure S5.