RNA-seq reveals global CELF2 regulation in T cells and sequence analysis suggests coregulation by the RBFOX family. (A) Venn diagram of the overlap of unique, high-confidence LSVs (Vaquero-Garcia et al. 2016) that showed significantly altered splicing (|ΔΨ| ≥ 20% with probability ≥95%) upon depletion of CELF2 by shRNA in unstimulated (left), stimulated (right), or both (middle) JSL1 Jurkat T cells (Fisher's exact (FE) test, two-tailed P < 7 × 10−192). (B) Scatterplot comparing E(ΔΨ) values for the most changing junction from the 271 unique, coregulated LSVs upon CELF2 depletion in unstimulated (X) and stimulated (Y) Jurkat T cells. (C) Fraction of high-confidence cassette exons derived from the union of events in panel A, containing splice site proximal CELF2 CLIP-seq peaks that were repressed by CELF2 (red, n = 265), enhanced by CELF2 (green, n = 371), or unresponsive (gray, n = 1668) (FE test, two-tailed). (D) Distribution of enrichment significance (−log10[P-value], hypergeometric test comparing regulated to unresponsive sets) for the occurrence of all pentamers upstream (left) or downstream (right) of CELF2-repressed exons. Top three pentamers for each are shown in the inset and motifs known to be bound by the CELF family or RBFOX family are highlighted in blue or orange, respectively. See Supplemental Table S2 for all values. (E) Cumulative distribution function (CDF) of the AVISPA predicted impact on splicing (normalized feature effect, NFE) (Barash et al. 2013) for downstream [U]GCAUG motifs for the subsets of events that contained it (Kolmogorov-Smirnov two-sample test). Fraction of each set with a downstream [U]GCAUG is given in inset with colors corresponding to the key (FE test, two-tailed). (F) CDF showing the average conservation (phastCons 46way, placental mammals) for the core GCAUG motif for the subsets of events that contained it downstream of the alternative exon (Kolmogorov-Smirnov [KS] two-sample test). Fraction of each set with a highly conserved downstream GCAUG (see Methods) is given in inset with colors corresponding to the key (FE test, two-tailed).
