Figure 5.

Misregulation of cassette exon splicing in c9ALS affects transcripts with roles in diverse molecular pathways. (a) Wiggle plots from RNAseq data and qRT-PCR bar graphs (mean ± s.e.m.) of top differentially misregulated cassette exons (FDR < 0.05, |dI| ≥ 0.1) in c9ALS cerebellum (see additional examples in Supplementary Fig. 8). Shown is an example of the 3 technical qRT-PCR replicates performed for each AS event in c9ALS (N=9) and sALS (N=10) and compared to controls (N=9). (b) qRT-PCR bar graphs of the same differentially misregulated cassette exons shown in a in the cerebellum of c9ALS (N=9) and sALS (N=10) compared to a disease control group (PSP: progressive supranuclear palsy, N=13). Relative mRNA levels shown in all bar graphs were normalized to the endogenous control, RPLP0, and respective controls (mean value set to 1). The full list of primers used in this study can be found in Supplementary Table 11.Statistical differences were calculated by one-way ANOVA with Bonferroni post-hoc test (*P < 0.05, **P < 0.01, ***P < 0.005, ^#P < 0.001). Exact P values can be found in Supplementary Fig. 8. (c) Gene-association network of the top most significant misregulated cassette exon events in c9ALS cerebellum (FDR < 0.05, |dI| ≥ 0.1). dl: differential index value. Genes are represented by nodes of different colors, which vary according to degree. The size of the node denotes neighborhood connectivity: nodes are bigger if they are connected to other nodes with higher connectivity. Edges are colored according to edge betweenness to indicate the proximity to other nodes, with low betweeness (closer proximity) meaning larger influence to other nodes. GO annotations for different interconnected cellular pathways are indicated. A more complete figure containing all gene names for each of the nodes can be found in Supplementary Figure 11.