Figure 4.

SA Functions through PP2A in Regulating Root Development

(A) SA treatment promoted the phosphorylation of PIN2 in Col-0 to a similar degree as that in pp2aa1-6. Roots of 7-day-old Col-0 and pp2aa1-6 seedlings were treated with DMSO or 40 μM SA for 60 min and were then sampled for protein isolation and western blot. The shifted bands indicate the phosphorylated PIN proteins (upper panel). Asterisk indicates a non-specific band that contributes partially to the signal. The same membrane was stripped and probed with a PIP2;1 antibody to indicate the loading (upper panel). Ponceau staining is shown in the bottom panel.

(B) Phosphorylation with ³²P-ATP revealed that SA treatment increased the phosphorylation of His-PIN2-HL in Col-0, whereas this increase was attenuated in pp2aa1-6. Upper panel: autoradiography is shown; lower panel: CBB is shown.

(C) Representative images revealing the hypersensitivity of pp2aa1-6 to SA. Col-0 and pp2aa1-6 seedlings were grown on plates with SA. Scale bars, 2 cm.

(D) pp2aa1-6 was hypersensitive to SA in root growth inhibition. Col-0 and pp2aa1-6 seedlings grew on plates with SA for 7 days and then the primary root length was measured. n = 11–28. p values were calculated by a two-tailed t test for indicated pairs of Col-0 and pp2aa1-6 at a certain concentration of SA.

(E and F) pp2aa1-6 was hypersensitive to SA in terms of interfering with root gravitropism. Col-0 (E) and pp2aa1-6 (F) seedlings grew on plates containing different concentrations of SA for 7 days, and the root tip angles were measured by ImageJ and shown as polar bar charts. p values were calculated by a two-tailed t test in (E) and (F) and indicate differences of variances by a further F-test in (F).

(G) The pp2aa1, a3 double mutant exhibited decreased sensitivity to SA. Col-0 and pp2aa1, a3 seedlings grew on plates with SA for 7 days and then the primary root length was measured. n = 11–25. p values were calculated by a two-tailed t test for indicated pairs of Col-0 and pp2aa1, a3 at the given concentration of SA.

(H) The pp2ac3, c4 double mutant exhibited decreased sensitivity to SA. Col-0 and pp2ac3, c4 seedlings grew on plates with SA for 7 days and then the primary root length was measured. n = 10–21. p values were calculated by a two-tailed t test for indicated pairs of Col-0 and pp2ac3, c4 at the given concentration of SA.

See also Figures S3 and S4.