Fig. 2.

Involvement of MKK5 in salt stress responses. RNA gel blot analysis of FSD2 and FSD3 transcript levels in response to salt stress in wild-type Arabidopsis plants, or the mkk2, mkk4, and mkk5 mutants. Total RNA was extracted from wild-type (WT) or mutant plants without stress treatment (control, C) or subjected to NaCl (150mM) for 4h (S). The actin gene ACT2 was used as a loading control. (B) Relative GUS activity driven by FSD2 or FSD3 promoter, respectively, showed the response of FSD2 and FSD3 to NaCl. The promoters of the FSD2 and FSD3 genes were fused to the GUS reporter gene and tested for their response to NaCl in transiently transformed protoplasts from wild-type plants or the mkk5 mutants. The MKK5 genes were cloned and co-transformed into protoplasts of the mkk5 mutant. Transformed protoplasts were pre-incubated 1h with 5 M PD98059 before treatment with 150mM NaCl. Protoplasts were isolated from wild-type (M) and the mkk5 mutant (m) leaves. (C) Relative GUS activity driven by FSD2 or FSD3 promoter, respectively, showed the response of FSD2 and FSD3 to NaCl. The promoters of the FSD2 and FSD3 genes were fused to the GUS reporter gene and tested for their response to NaCl in transiently transformed protoplasts from wild-type plants or the mkk2 and mkk4 mutants. The MKK2 and MKK4 genes were cloned and co-transformed into protoplasts of the mkk2 or mkk4 mutant, respectively. Protoplasts were isolated from wild-type (M) and mutant (m) leaves. All experiments were repeated at least three times with similar results.