Extended Data Fig. 9: Analysis of somatic cancer mutations in RING-domain E3 ubiquitin ligases.
From: Mechanical regulation of glycolysis via cytoskeleton architecture
a, Cancer mutations in TRIM21 RING domain found in proximity to conserved cysteines and histidines. b, Quantification of the number of genes that displayed one or more mutation at positions C1–C8 of the RING domain. c, Summary table of missense and nonsense mutations found in 118 genes at the conserved cysteine and histidine positions C1–C8. *Stop codon. d, Distribution of mutations across C1–C8. e, Frequency of missense and nonsense mutations at positions C1–C8, and overall. f, TRIM21–GFP or TRIM21(C54Y)–GFP expression in HBEC76 cells, counterstained for F-actin with phalloidin. Scale bar, 10 μm. Representative data from a single imaging experiment. g, Representative images of H2009 cells expressing TRIM21–GFP with indicated cysteine mutations, counterstained for F-actin with phalloidin. Scale bar, 10 μm. The experiment was performed once. h, Representative images of H2009 cells expressing GFP, TRIM21–GFP or GFP-tagged TRIM21 with cancer-relevant mutations, counterstained for F-actin with phalloidin. Scale bar, 10 μm. The experiment was performed once. Mutations shown in g cause protein aggregation, whereas non-cysteine mutations in h do not. i, Abundance of PFKP on stiff and soft substrates when HBEC76 cells expressed TRIM21–GFP or TRIM21(C54Y)–GFP. The experiment was performed once. j, Effect of expressing TRIM21 or TRIM21(C54Y) on glycolytic rates of HBEC76 cells, normalized to cell number. Data are from three independent experiments, shown as mean glycolytic rate ± s.e.m. Dotted line, glycolytic rates of HBEC76 cells on stiff substrates without overexpressing TRIM21, as indicated in Fig. 1c. Protein abundance was normalized to GAPDH (i).
