Figure 4.

Novel atg2 and atg18a alleles recovered as lon2 suppressors. (a) The L60 suppressor was backcrossed to the original lon2-2 line, and genomic DNA from pooled IBA-sensitive F₂ seedlings was sequenced and analyzed as in the legend to Figure 1(d). (b) ATG2 gene diagram with boxes and lines representing protein-coding regions and introns, respectively. The positions of new atg2 mutations identified as lon2 suppressors are shown in green, previously described EMS-derived lon2 suppressors [48] are in gray, and a T-DNA insertion allele [64] is indicated by a triangle. The partial alignment shows predicted Brassicaceae ATG2 proteins, including 2 A. thaliana ATG2 splice variants predicted by the latest genome annotation (Araport 11); the alternative ATG2 splice variant is interrupted by the atg2-6 mutation. aa, amino acids. (c) ATG18a gene diagram showing atg18a mutations identified as lon2 suppressors (teal) and a previously described T-DNA insertion allele [65] (triangle). (d) Lateral root density of 8-day-old wild type (WT), lon2-2, atg2-4, lon2-2 atg2, and lon2-2 atg18a seedlings grown without or with IBA. Error bars show standard deviations (n = 8). Statistically significant (P < 0.0001) differences determined by one-way ANOVA are depicted by different letters above the bars. (e) Extracts from 6-day-old seedlings were processed for immunoblotting with antibodies to the indicated proteins.