Fig. 3: Characterization of alternative splicing events in THOC6 affected hNPCs.
A Differentiation protocol to derive human neural progenitor cells from affected and unaffected ESC/iPSCs. B Combined rMATS summary results of statistically significant AS events in THOC6W100*/W100* and THOC6E188K/E188K hNPCs relative to THOC6W100*/+ control hNPCs. Event type (pie chart) and inclusion status (bar chart). Yellow, higher inclusion in affected condition. Blue, higher inclusion in unaffected conditions. C Significant splice site strength score differences at mis-spliced events in affected hNPCs based on the maximum entropy model. Black, non-significant events. Yellow, higher inclusion in affected condition. Blue, higher inclusion in unaffected conditions. Significance obtained by two-tailed, unpaired t test. **** indicates p = <0.0001. Transcript number per AS gene D and AS event length E in THOC6W100*/W100 and THOC6E188K/E188K vs. THOC6W100*/+ NPCs. Black, non-significant events. Yellow, higher inclusion in affected condition. Blue, higher inclusion in unaffected conditions. Significance obtained by two-tailed, unpaired t test. **** indicates p = <0.0001. F RT-PCR AS validations of SE (ABAC1, POU2F2) and RI (MAPK15) events in n = 3 additional biological replicates of hNPCs per genotype (6 per condition) with quantified mis-spliced ratios. Significant outliers were identified for POU2F2 and were then excluded from analysis. Data shown as mean ± SEM. Significance obtained by two-tailed, unpaired t test. UNAFF, unaffected heterozygous control (black); AFF, homozygous affected (red). G Venn diagram of overlap of THOC6 W100*/W100* and THOC6E188K/E188K AS genes and all syndromic intellectual disability genes included in the SysID database. Overlap significance tested by one-sided Fisher’s exact test. ASG, alternatively spliced genes. H Metascape analysis on combined significant mis-spliced events (FDR < 0.05) in THOC6E188K/E188K and THOC6W100*/W100* NPCs.
