Extended Data Fig. 7: Autophagic phenotype analysis of vsr mutants.
(a) Photographs of the seedlings from the indicated genotypes after maintaining them for 7 d in complete darkness. Seedlings were grown 7 d on half-strength MS under long-day conditions before transfer to dark. Note that starvation-induced chlorosis is enhanced in vsr5vsr6vsr7 or vsr1vsr6vsr7, compared with WT or complemented lines (GFP-VSR7/vsr5vsr6vsr7 and GFP-VSR1/vsr1vsr6vsr7). Scale bar, 1 cm. (b) Quantification analysis of the total chlorophyll content of seedlings in (a). Data are presented as means ± SD of three independent experiments. (c) Quantification analysis of the NBR1 proteins level upon infection with Pto DC3000 avrRpm1. The immunoblot intensity was normalized by the loading control of anti-actin, and the first lane (0 h) in each experiment was arbitrarily set to 1. Data are presented as means ± SD from three independent experiments. The differences were compared among WT and vsr mutants at the indicated time points. (d) Immunoblot analysis of NBR1 accumulation upon infection with Pto DC3000 avrRpm1. Total proteins were extracted from 5-day-old WT, vsr5vsr6vsr7, vsr5vsr6vsr7atg5, and atg5 seedlings at the indicated time points (0 and 6 h), followed by immunoblot analysis with anti-NBR1 antibody. The cytoplasmic marker anti-cFBPase is used as a loading control. The immunoblot intensity was normalized by the loading control of anti-cFBPase, and the first lane (WT, 0 h) was arbitrarily set to 1. Quantification analysis of the NBR1 proteins level is shown on Right. Data are presented as means ± SD from three independent experiments. (e) Quantification analysis of the ratio of free eYFP to eYFP-ATG8e proteins level upon infection with Pto DC3000 avrRpm1. Data are presented as means ± SD from three independent experiments. Different letters above bars indicate a significant difference at the P < 0.05 level by one-way ANOVA with Fisher’s LSD (b, d, and e) or Tukey’s (c) multiple comparisons test. Exact P-values of statistic tests in (b-e) are provided in the Source data file.
