Mitogen-activated protein kinases 3 and 6 are required for full priming of stress responses in Arabidopsis thaliana - PubMed (original) (raw)

Attenuation of BTH-IR, SAR, and Infection-Induced Dual TEY Phosphorylation in mpk Mutants. (A) Reduction of BTH-IR. Wild-type, mpk3, and mpk6 plants were pretreated with wettable powder carrier (−) or 100 μM BTH (+). Three days later, leaves of the plants were dip inoculated with a high titer of Pst DC3000 (5 × 108 cfu mL−1). Four days after infection, leaf discs were harvested and analyzed for the amount of bacteria, while another portion of infected leaves was examined for disease symptoms. Note that in some repeats of the experiment, the BTH-IR was completely abolished in mpk3. The experiment was done seven times with similar results. The values shown are means +

sd

(n = 10 biological replicates). Asterisks indicate significant differences (Student's t test, n = 10, P < 0.05). (B) Attenuation of SAR. Three lower leaves of wild-type, mpk3, and mpk6 plants were infiltrated with a suspension (5 × 105 cfu mL−1) of avirulent Pst DC3000 harboring the avirulence gene avrRpt2 (+). The bacteria were suspended in 10 mM MgCl2. Control plants were infiltrated with MgCl2 in the absence of bacteria (−). Three days later, two upper leaves were challenge-infected with a suspension of Pst DC3000 (5 × 105 cfu mL−1) in MgCl2. Leaf discs were harvested from challenge-infected leaves and analyzed for the amount of bacteria after 3 d. The experiment was performed three times with similar results. The values shown are means +

sd

(n = 8 biological replicates). Asterisks indicate significant differences (Student's t test, n = 8, P < 0.05). (C) Reduction of induced dual TEY motif phosphorylation. Same experimental setup as in (B) but harvesting aliquots of leaves 2 h after challenge infection with Pst DC3000 to analyze dual phosphorylation of the TEY motif by SDS-PAGE, protein gel blotting (WB), and immunodetection with polyclonal antibodies. Prior to immunodetection, the blot was stained with Ponceau S to assess whether loading was equal. The experiment was done three times with similar results. Note the absence of pTEpY immunodetection bands in the mpk3 and mpk6 mutant. This data confirms that the immunoreactive material detected by the pTEpY antibody is phosphorylated MPK3 and MPK6. For quantification of immunodetection signals, see Supplemental Figure 1 online.