Journal: bioRxiv
Article Title: Distinct functions of PBRM1 and BAP1 reconcile the course of kidney cancer evolution and disease progression
doi: 10.64898/2026.04.30.721837
Figure Lengend Snippet: (A) Schematic of CRISPR/Cas9-mediated BAP1 knockout (BKO) in patient-derived normal (PDON) and tumour (PDOT) organoids. The cross indicates unsuccessful expansion. (B) Tracking of variant allelic frequency (VAF) in normal kidney organoids following electroporation with guide RNAs targeting PBRM1 or BAP1 . (C) Schematic of CRISPR/Cas9-mediated BAP1 knockout (BKO) in normal organoids (K1088N1_GN1) with concurrent CDKN2A and VHL loss (CV background). (D) Principal-component analysis (PCA) of bulk transcriptomes from parental and BAP1 -engineered organoid lines. (E) GSEA of BAP1 -KO tumour organoids versus parental controls using the Hallmark E2F Targets gene set. (F) Comparison of normalised expression (DESeq2 VST) of selected genes between BAP1 -KO and parental tumour organoids ( n = 5 PDOT, n = 6 PDOT-BKO). Unpaired t -test; *: p < 0.01. (G) GSEA of BAP1 -KO tumour organoids versus parental controls using the Gene Ontology Biological Process (GOBP) Chromosome Segregation gene set. (H) Representative DAPI staining showing micronucleus presence in BAP1 -KO and parental tumour organoids. Scale bar, 50 µm. (I) Violin plot showing the number of clonal copy-number variants (CNVs), excluding chromosome 3p loss, in tumours with clonal BAP1 or PBRM1 mutations. Unpaired t -test; p = 0.0703. Clonal PBRM1 , n = 25; clonal BAP1 , n = 7. (J) Heatmap showing normalised accessibility scores for the top 1000 differentially accessible regions (DARs) between BAP1 -WT and BAP1 -KO organoids. (K) Genomic annotation of regions with increased (top) or decreased (bottom) chromatin accessibility following BAP1 loss. UTR, untranslated region. (L) Volcano plot showing differential transcription-factor binding activities between BAP1 -WT and BAP1 -KO tumour organoids. Significance was defined by −log10(p-value) above the 95% quantile and differential binding scores in top 5% in each direction. See also Figure S8 and Table S7.
Article Snippet: A total of one million single cells were pelleted and resuspended in 100 μL electroporation buffer (Amaxa P3 Primary Cell Solution, Lonza), prepared by adding 18 μL of Supplement to 82 μL P3 Solution.
Techniques: CRISPR, Knock-Out, Derivative Assay, Variant Assay, Electroporation, Comparison, Expressing, Staining, Binding Assay
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