Fig. 5. LLG1 regulates mekk1 cell death.
a The llg1 mutants suppress growth defects triggered by silencing MEKK1. The plant images were photographed at 3-weeks after inoculation with Agrobacterium carrying the indicated VIGS vectors. Ctrl is the vector containing GFP. Scale bar, 1 cm. b The llg1 mutants suppress cell death and H2O2 accumulation induced by silencing MEKK1. The leaves from plants in a were stained by trypan blue for cell death and DAB for H2O2 accumulation. Scale bar, 0.5 cm. c Expression of HA-LLG1 in llg1-2 restores the cell death triggered by silencing MEKK1. #1 and #2 are two representative pLLG1::HA-LLG1 transgenic lines in llg1-2. Scale bar, 0.5 cm. d Protein expression of HA-LLG1 in pLLG1::HA-LLG1/llg1-2 transgenic plants. e The llg1-1mekk1 mutant enhances growth defects of mekk1. The seedlings grown on ½MS plate at 22 °C were photographed at 2-weeks post-germination. Scale bar, 0.5 cm. f The fresh weight of llg1-1mekk1 is less than mekk1. The data are shown as mean ± SE (n = 3). P = 3.63 × 10−5 (column 3 and 5). The asterisk indicates statistical significance by using two-sided two-tailed Student’s t test (***P < 0.001). g llg1-1 mutant enhances the expression of PR1 in mekk1. The expression of PR1 was determined with the plants in e and normalized to the expression of UBQ10. The data are shown as the mean ± SE of four biological repeats (n = 4). P = 1.04 × 10−7 (column 3 and 4). The different letters indicate the significant difference determined by one-way ANOVA followed by the Tukey test (P < 0.05). h The fer-4mekk1 mutant enhances growth defects of mekk1. The seedlings grown on ½MS plate at 22 °C were photographed at 2-weeks post-germination. Scale bar, 1 cm. i The fresh weight of fer-4mekk1 mutant is less than mekk1. The data are shown as mean ± SE (n = 3). P = 6.92 × 10−4 (column 3 and 4). The asterisk indicates statistical significance by using two-sided two-tailed Student’s t test (***P < 0.001). j fer-4 mutant enhances the expression of PR1 in mekk1. The expression of PR1 was normalized to the expression of UBQ10 and the data are shown as the mean ± SE of four biological repeats (n = 4). P = 1.87 × 10−6 (column 3 and 4). The different letters indicate the significant difference determined by one-way ANOVA followed by the Tukey test (P < 0.05). The assay was performed as in g. k High temperature did not alleviate fer-4mekk1 growth defects. The seedlings grown on ½MS plate at 28 °C were photographed at 2-weeks post-germination. Scale bar, 1 cm. l The fresh weight of fer-4mekk1 mutant is less than mekk1 at 28 °C. The seedlings in k were used for measuring fresh weight. The data are shown as mean ± SE (n = 3). P = 1.81 × 10−6 (column 3 and 4). The asterisk indicates statistical significance by using two-sided two-tailed Student’s t test (***P < 0.001). To measure the weight of mekk1, llg1mekk1, and fer-4mekk1 mutants, 10 plants were pooled and the weight of individual plants was averaged. The above experiments were repeated 3–4 times with similar results.