Fig. 3: Assessing splicing factor activity in complex molecular contexts.

a Indisulam promotes protein-level degradation of splicing factor RBM39, inducing significant changes in the exon inclusion repertoire of the cell. b Differential proteomic expression of splicing factors. Relationship between fold change LFQ values comparing Indisulam-treated (5 μM, 6 h) and untreated samples, and p-values obtained from a two-sided T-test in IMR-32 cells. Red, RBM39. c Ranking of splicing factors according to their estimated protein activities upon treatment with Indisulam (5 μM, 6 h) in IMR-32 cells. Red, RBM39. d Distributions of estimated protein activities upon combinatorial perturbation of splicing factors. Red, corresponding knocked down genes in each case. Top, experimental outline. Studies analyzed include PRJNA223244³¹, PRJNA498529³⁰, PRJNA587741²⁹, and PRJNA321560²⁸. e The SF3b complex is composed of 8 proteins. When perturbing spliceosome assembly with drugs, we hypothesize that proteins in the SF3b complex and proteins interacting with the complex should decrease their activity. Adapted from Rogalska et al.⁸. f Number of proteins at each shortest path length to reach any protein of the SF3b complex in the STRINGDB protein-protein interaction network. Color, whether genes belong to the U2snRNP. g Top, mechanisms of action of splicing-targeting drugs: H3B-8800 and Pladienolide B bind the SF3b complex; E7107 prevents the binding of U2 complex to RNA; Isoginkgetin and Spliceostatin A impede the transition from A to B complex. Bottom, distributions of splicing factor activities upon perturbation with different splicing-targeting drugs, considering the shortest path lengths of each splicing factor to the SF3b complex. Color, whether genes belong to the U2snRNP. Studies analyzed include PRJNA371421³³, PRJNA685790⁶⁷, PRJNA662572⁶⁸, PRJNA380104⁶⁹, PRJNA354957⁷⁰, and PRJNA292827⁷¹. In box and whiskers plots in the panel, the median is marked by a horizontal line, with the first and third quartiles as box edges. Whiskers extend up to 1.5 times the interquartile range, and individual outliers are plotted beyond. The number of proteins per path-length bin matches panel (f). Source data are provided as a Source Data file.