Dysregulated splicing in breast cancer is enriched for snRNA-responsive splice sites and exons. (A) snRNA-responsive intron excision within Lck interacting transmembrane adaptor 1 (LIME1). Dashed box indicates differentially retained intron, for which U1 snRNA depletion promotes intron retention. (Top rows) RNA-seq read coverage from control and U1-depleted MCF-7 cells. (Bottom rows) Read coverage from the 10 breast cancer biopsies with the highest and lowest U1 levels. (B) Distribution of Ψ values across breast cancer specimens (N = 136), stratified by U1 snRNA KD-sensitive (red) and -insensitive (gray) splicing in MCF-7 cells. Dashed lines indicate splicing events for which the alternative spliced sequence is preferentially excluded from the mature transcript following U1 snRNA KD in MCF-7 cells. Solid lines indicate events for which the alternatively spliced sequence is preferentially included following U1 snRNA KD. For retained introns, the dashed line indicates intron retention. Plots are restricted to exons and introns that were annotated as alternatively spliced (see Methods). Note that events that are uniformly excluded or included from the mature mRNAs across the cohort have Ψ values close to zero and one, respectively, whereas intermediate Ψ values indicate heterogeneous splicing that varies within or across patients. (C) Proportions of snRNA-dependent and snRNA-independent events that are differentially spliced in at least 10 primary breast cancer biopsies relative to patient-matched peritumoral normal samples (N = 107). Samples are from the TCGA breast cancer cohort. Red indicates U1; green, U2; blue, U4; purple, U6. (D) Cumulative density functions comparing differential splicing of U1 snRNA KD-sensitive (red) and -insensitive (gray) splicing events across breast cancer patients, as in C. The plots illustrate the percentage of primary breast cancer biopsies showing aberrant splicing relative to their patient-matched normal sample, across the TCGA cohort. Splicing events were stratified as U1 snRNA KD-sensitive (red) and -insensitive (gray) based upon the observed splicing in MCF-7 cells. (N) Number of events that could be reliably quantified in the U1 snRNA KD sample. Statistical significance was calculated using the Kolmogorov–Smirnov test.
