Fig. 5.
Induced expression of AvrRps4 in Arabidopsis leads to ICS1 and PR1 expression, but not MAPK activation. (A and B) ICS1 (A) and PR1 (B) expression after induction with E2 for 2, 4, and 8 h in SETI (left panel), SETI_KRVYmut (middle panel), and SETI_eds1 (right panel) leaf samples. Five-week-old SETI and SETI_KRVYmut leaves were infiltrated with 50 μM E2. Samples were collected at 0, 2, 4, and 8 hpi for RNA extraction and subsequent qPCR. The expression level is presented as relative to EF1α expression. Each data point represents one technical replicate. The black line represents the mean of the technical replicates. This experiment was repeated three times independently with similar results. Significant differences relative to the untreated samples were calculated with t-test, and the P-values are indicated as ns (non-significant), P>0.05; *P<0.05; **P<0.01; ***P<0.001. (C) Activation of MAPKs in Col-0, SETI_WT, SETI_KRVYmut, and SETI_eds1 seedlings by E2 induction of effector AvrRps4 or mutant AvrRps4KRVY-AAAA. Seedlings grown in liquid culture at 7 DAG were treated with 50 μM E2 for the indicated times (0, 2, 4, 6, or 8 h) and samples were collected. Col-0, SETI_WT, SETI_KRVYmut, and SETI_eds1 seedlings treated with 100 nM flg22 for 10 min (10 min*) were used as positive control. Proteins were extracted from these seedlings and phosphorylated MAPKs were detected using p-p42/44 antibodies. Arrowheads indicate phosphorylated MAPKs (black, pMPK6; grey, pMPK3; white, pMPK4/11). Ponceau staining was used as loading control. (This figure is available in colour at JXB online.)