Figure 3.

Transcriptome-Wide Comparison of Splicing Variant Patterns between Control and NMD-Impaired Arabidopsis Seedlings.

(A) Size-proportional Venn diagram of significantly altered AS events (with numbers of corresponding genes in parentheses) comparing lba1 upf3-1 and CHX treatment (FDR ≤ 0.1) and lba1 upf3-1 with the two single mutants (double mutant FDR ≤ 0.1; single mutant P ≤ 0.1). Asterisks provide information on statistical significance, and contradictory events with significant changes in opposite directions have been excluded (for numbers, see Supplemental Table 1 online).

(B) Relative frequencies of AS types in all detected events as well as for the indicated combinations of NMD impairments: lba1 upf3-1 + CHX (1), changed under both conditions; lba1 upf3-1 + SM – CHX (2), changed in lba1 upf3-1 and in at least one of the single mutants but not upon CHX treatment; all NMD events (3), combination of the two previous sets. alt 5′/3′ ss, Alternative 5′/3′ splice sites.

(C) Direction of splicing change for subsets 1, 2, and 3 defined in (B). Direction of change for alternative 5′/3′ splice sites refers to usage of the downstream and upstream splice sites, respectively.

(D) Gene models of alternatively spliced regions (top), representative coverage plots (relatively scaled to the same height in all figures) with the altered region in black (middle), and quantitative analysis of splicing variants (bottom) for selected AS events identified by RNA-seq. Splicing variant ratios were determined by Bioanalyzer quantitation (mean values + sd, n = 3). Boxes and lines in gene models represent exons and introns, respectively, with black boxes depicting coding regions (according to the representative gene model annotated at TAIR10; SI). Asterisks indicate translation termination codons, and arrowheads illustrate the binding positions of primers used in RT-PCR. WT, Wild type. Bars = 100 nucleotides.