Protein-protein interactions in the RPS4/RRS1 immune receptor complex
Fig 5
PAD4 attenuates EDS1/AvrRps4 association.
(A) The EDS1/PAD4 complex strongly reduced EDS1/AvrRps4 co-immunoprecipitation in planta. EDS1-Myc or EDS1-Myc/PAD4-HA were transiently co-expressed with AvrRps4-GFP, AvrRps4E187A-GFP or GFP in N. benthamiana leaves. Immunoprecipitations were performed using anti-Myc agarose beads and then analyzed by immunoblot with the indicated antibodies. AvrRps4C represents processed AvrRps4 C-terminus. (B) The EDS1/SAG101 complex can associate with AvrRps4. HA-EDS1 or HA-EDS1 with SAG101-Myc were transiently co-expressed with AvrRps4-GFP or GFP in N. benthamiana leaves. Immunoprecipitation was performed using anti-HA agarose beads and then analyzed by immunoblot with the indicated antibodies. (C) BiFC analysis reveals that EDS1/AvrRps4 interaction is reduced in the presence of PAD4-HA but not of SAG101. Cytoplasmic aggregations are reduced in the presence of PAD4. BiFC assays were performed by co-expression of the indicated proteins in N. benthamiana. Images were obtained at 2 dpi. The experiment was repeated three times with similar results. Scale bar = 10 μm.