Fig. 2.

(A) Five- to 6-wk-old WT leaves were syringe infiltrated with water (−) or flg22 (+) and mRNA levels of known TGS factors (NRPD2, NRPD7, AGO4, AGO6, NRPE7, NRPE5, IDN2, KTF1, DRD1, and MET1) were monitored at 3 and 6 h post-flg22 treatment with SD as in Fig. 1, Left. FRK1 and PR1 transcript levels were also analyzed at 3, 6, and 9 h posttreatment with SD as in Fig. 1, Right. Similar results were obtained in five independent experiments. (B) GUS mRNA levels were monitored as in A from AGO4p:GUS-elicited plants with SD as in Fig. 1. Similar results were obtained in two independent experiments. (C) WT plants were treated as in A for 9 h and AGO4, NRPE5, and ACTIN8 protein levels monitored by Western blot analyses. Similar results were obtained in two independent experiments. (D) WT plants were treated as in A and ROS1 transcript levels were analyzed at 3, 6, and 9 h posttreatment with SD as in Fig. 1. Similar results were obtained in three independent experiments. (E) Methylation levels at AtSN1 analyzed by bisulfite sequencing in plants treated as in A. The region analyzed contains four CG, seven CHG, and 33 CHH. Asterisks represent significant differences (**P < 0.01). Similar results were obtained in five independent experiments, with kinetics of DNA demethylation starting at 6 or 9 h after flg22 treatment from one experiment to the other. (F) mRNA levels of AtSN1 treated as in A in WT and ros1–4 leaves with SD as in Fig. 1. Similar results were obtained in two independent experiments. Of note, the kinetics of DNA demethylation depicted in E are not directly comparable with the kinetics of AtSN1 induction presented in F. These experiments are independent. (G) Methylation levels at AtSN1 in plants treated as in A for 6 h and analyzed as in E. Asterisks represent significant differences (**P < 0.01). Similar results were obtained in two independent experiments, with a kinetics of DNA demethylation occurring at 6 h in one experiment and 9 h in the other.