Fig. 5

Aberrant SFPQ intron retention in diverse genetic forms of ALS and its interplay with the SFPQ protein. a Left, genome browser views of RNA-seq profiles for the intron-retaining gene SFPQ in control and VCP^mu samples at iPSC, NPC and pMN stages. 9 kb intron 9/9 of interest is indicated with yellow box. Right, bar graphs quantifying percentage IR across the entire time course in control and VCP^mu samples (mean ± s.d.; Fisher count test). b Bar plots displaying SFPQ IR levels measured by qPCR; white bars = controls, black bars = VCP^mu. IR levels at each timepoint were compared with that of the iPSC stage of the same group. N = 3 control lines and N = 4 VCP lines (mean + s.d. *p < 0.05, **p < 0.01, one-way ANOVA with Dunnet correction for multiple comparisons). c Bar plots showing the percentage SFPQ IR for FUS^mu or SOD1^mu MNs in control compared with (mean ± s.d.; Fisher count test). d Bar plots depicting the level of enrichment in RBP-binding to the retained intron compared with the non-retained introns within the same gene; blue bars = SFPQ gene, green bars = FUS gene. e–f Genome browser view of SFPQ eCLIP crosslinking events along SFPQ and FUS transcript annotations. Grey boxes highlight the location of retained introns. g Bar plots showing the level of nuclear and cytoplasmic localisation of IR transcripts measured by qPCR. IR levels in each fraction were compared with that of the nuclear fraction of control iPSCs. N = 3 control lines and N = 4 VCP lines, mean + s.d. P-value from two-way ANOVA with Tukey correction for multiple comparisons. h Subcellular localisation of SPFQ determined by immunocytochemistry in iPSCs, NPCs and pMNs. The ratio of the average intensity of the SFPQ staining in the nucleus (N) vs cytoplasm (C) was automatically determined in both CTRL and VCP^mu cells. Data shown is average N/C ratio (±s.d.) per field of view from four control and four VCP^mu lines. P-value from unpaired t-test with Welch’s correction. See also Supplementary Fig. 7d