Extended Data Fig. 9: The proteoglycan VCAN modestly increases resistance to ferroptosis and lipoprotein uptake in cancer cells.

a. Schematic of CRISPR screens in 786-O (ccRCC) and Karpas299 (lymphoma) cells transduced with a proteoglycan-focused sgRNA library (55 genes). 786-O cells were subjected to a proliferation-based screen in the presence or absence of the GPX4 inhibitor, ML162 (left). Karpas299 cells were subjected to flow cytometry-based cell sorting for high and low fluorescent populations after DiI-LDL treatment. b. Differential gene scores from the 786-O CRISPR screen under ML162 treatment relative to untreated cells. Negative scores indicate genes whose loss sensitizes cells to ML162. UGDH served as a positive control. c. Differential gene scores in high DiI-LDL uptake versus low uptake CRISPR screen in Karpas299 cells. Negative scores indicate genes whose loss reduces LDL uptake. UGDH served as a positive control. d. Quadrant map showing overlapping hits from both screens; genes essential in both are highlighted, with VCAN emerging as a shared hit. e. Sanger sequencing of VCAN gene exon 9 in Karpas299 parental cells (WT) or transduced with sgVCAN. f. Proliferation (log₂ doublings, 5 days) of 786-O transduced with a sgControl or sgVCAN under the indicated concentrations of ML162. g. Proliferation (log₂ doublings, 5 days) of Karpas299 cells transduced with a sgControl or sgVCAN under the indicated concentrations of ML162. h. Cellular uptake of DiI-LDL in the indicated Karpas299 cells transduced with a sgControl or sgVCAN, quantified as median PE intensity. i. Tumour weights resulting from implantation of Karpas299 cells transduced with a sgControl or sgVCAN in mice. f-h: Bars represent mean ± s.d.; i: Boxes represent median, first and third quartiles, and whiskers are range. f-h, n = 3 biological replicates; i, n = 10 biological replicates. Statistics by two-sided unpaired t-tests compared to sgControl-expressing cells (f-i).

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