Genome-wide CRISPR screen identifies splicing factor SF3B4 in driving hepatocellular carcinoma

Although genome sequencings have recognized many cancer-associated genes in hepatocellular carcinoma (HCC), distinguishing their functional effect remains challenging. Leveraging on a genome-wide CRISPR knockout (KO) screening, we uncovered spliceosome factors as major survival essential genes in HCC and up-regulations of ferroptosis suppressors [particularly glutamate-cysteine ligase catalytic subunit (GCLC)] in lenvatinib resistance. Our KO screen in patient-derived HCC organoid showed splicing factor 3b subunit 4 (SF3B4) to be top-ranked, conferring prosurvival signal in HCC organoid and driving tumorigenic potentials in both hepatic progenitor organoids and hydrodynamic tail vein injection HCC murine model. The combined RNA immunoprecipitation sequencing, long-read isoform sequencing, and transcriptome revealed characteristic splicing landscape regulated by SF3B4 and identified T-box transcription factor 3 (TBX3) variant TBX3+2a as a potent downstream effector. Our findings highlighted vital roles of SF3B4 in HCC cell survival and tumor progression, and the phenomenon of ferroptosis resistance in patients unresponsive to first-line agent lenvatinib.