A combined enteric neuron-gastric tumor organoid reveals metabolic vulnerabilities in gastric cancer

Data S1. Focused library CRISPR screen quality and the aberrant splicing events detected upon knockout of the synthetic lethal gene hit MAGOHB in ARHGAP fusion-containing organoids, related to Figure 2.
A, More than 98% of the sgRNAs were detected within a 5-fold range from the mean reads of sgRNAs as highlighted by the shaded areas.
B, Screen data reproducibility between two biological replicates in the focused library screening.
C, Growth competition assay of organoid lines with (GX052-TO and GX121-TO) and without (GX049-TO, GX068-TO, GX069-TO and GX076-TO) ARHGAP fusion gene with MAGOHB knockout. n = 3.
D, GFP disruption percentage of organoids with MAGOHB knockout in terms of doubling frequency (Number of hours post-sgRNA transduction/Doubling time) normalized to safe harbor.
E, RNA-seq and splicing analyses revealed increased alternative splicing events, along with aberrant changes in spliced intron count at the fusion gene upon MAGOHB knockout, indicate its regulatory role in the splicing of CTNND1-ARHGAP26. Number of total differential alternative splicing events including alternative 3’ splice sites (A3SS), alternative 5’ splice sites (A5SS), mutually exclusive exons (MXE), retained introns (RI) and exon skipping (SE) in GX058-TO upon MAGOHB knockout (left panel). Number of differential alternative splicing events identified in CTNND1 and ARHGAP26 sequences in GX058-TO with MAGOHB knockout (right panel).
F, Fold change of spliced intron counts per million between GX058-TO control and its respective CTNND1-ARHGAP26 knockout clone (upper panel) or with MAGOHB knockout (lower panel) at the indicated genomic coordinates. The fusion gene is identified at chr11: 57810223:+chr5: 143014080 by STAR-Fusion. n = 2.
G, Relative fusion fragments per million total RNA-seq fragments (FFPM) of fusion gene knockout and MAGOHB knockout to controls analyzed by STAR-Fusion. n = 2.
H, Knockout of MAGOHB introduced reduction in CTNND1-ARHGAP26 fusion protein expression. The modest decrease in the fusion exon transcript revealed by STAR-Fusion when MAGOHB was knocked out is evident in the reduction of the fusion protein.
I, Enriched pathways for genes with at least 5 total differential alterative splicing events in GX058-TO with MAGOHB knockout using Hallmark gene sets. n = 2.
J, Enriched pathways of the downregulated genes in MAGOHB knockout organoids compared with control with reference to Hallmark gene sets. n = 2. The growth inhibition brought by MAGOHB knockout could be due to the reduction of the disease fusion protein level and/or the concomitant alternations of gene expressions in additional pathways.