A bacterial effector protein prevents MAPK-mediated phosphorylation of SGT1 to suppress plant immunity
Fig 2
RipAC associates with SGT1 in plant cells.
(A) CoIP to determine interactions between RipAC and SGT1s transiently expressed in Nicotiana benthamiana. The signal in the interaction between RipAC and AtSGT1a was weak and is shown with longer exposure. (B) Split-LUC assay to determine direct interaction between RipAC and SGT1 transiently expressed in N. benthamiana. (C) Pull-down assay to determine direct interaction between RipAC and SGT1a/b. His-tagged RipAC was incubated with immobilized GST-AtSGT1a/AtSGT1b/GUS. After 4 rounds of wash, bound proteins were eluted and subjected to western blot analysis using anti-RipAC antibody. Coomassie blue (CBB) staining showed the visualization of both input and GST pull-down proteins. (D) CoIP to determine interactions between RipAC and different truncated versions of NbSGT1 in N. benthamiana. The diagram summarizes the different domains of SGT1: TPR, N-terminal tetratricopeptide repeat (TPR) domain; CS, CHORD-SGT1 domain; SGS, C-terminal SGT1-specific domain; VR, variable region. The experiments in (A-C) were repeated at least 3 times with similar results. The experiment in (D) was repeated 2 times with similar results. In western blot assays, protein marker sizes are provided for reference. In (A) and (D), blots were stained with Coomassie Brilliant Blue (CBB) to verify equal loading.