Fig. 6

SHQ1 depletion affects pre-mRNA splicing efficiency. HPB-ALL or KOPTK1 cells were infected with lentiviruses expressing control shRNA (Ctrl) or SHQ1 shRNA (sh1 or sh2). a H/ACA snoRNA scaU93, base paired with U2 snRNA and guiding its pseudouridylation, in HPB-ALL cells expressing control shRNA (blue), SHQ1 shRNA-1(red), or shRNA-2(purple) was analyzed by qPCR. b Schematic of intron retention (IR) analysis. Exon–intron reads (a) relative to exon–exon reads (b) are defined as IR ratio which reflects the magnitude of pre-mRNA accumulation. c Empirical cumulative distribution of IR coefficients for 6809 genes in HPB-ALL cells. Curves represent IR ratio in control (black) and shRNA-2 mediated SHQ1 depletion (red). A rightward shift in the curve representing SHQ1-depleted cells indicates increased IR, namely increased pre-mRNA accumulation upon SHQ1 knockdown. d Genome-wide splicing and gene expression were analyzed in HPB-ALL and KOPTK1 cells. Numbers of genes with altered IR ratio were plotted in red at the pie chart, and those with no intron retention (NIR) were marked in black. Genes with IR ratios (SHQ1 knockdown vs control) more than 1.15 are considered aberrant intron retention. Heatmaps of mature mRNA expression from intron-retained genes are presented at the bottom (red indicates increased and blue decreased). Top genes with most decreased mature mRNA associated with increased IR ratios in SHQ1-deficient cells are highlighted in the box. e, f Validation of candidate genes shown in the box from d by assessing IR ratio, mature mRNA (e), and protein expression (f) in HPB-ALL cells. g Analysis of T-ALL associated oncogene expression in HPB-ALL cells by qPCR. In e and g, blue, red, or purple bars represent indicated RNA quantifications in control shRNA, SHQ1 shRNA-1, or -2-expressing cells. Data shown represent the means (±SEM) of three biological replicates; *p < 0.05, **p < 0.01, ***p < 0.001, unpaired t-test (a, e, g) or Kolmogorov–Smirnov test (c)