Fig 1: Alteration of tRF-3 m1A levels and tRF-3 targets in bladder cancer.a TRMT6 expression in TCGA BLCA (n = 404) compared to the normal (n = 28, TCGA and GTEX). Expression (Y-axis) is derived from RNA-seq on Log2 scale. Data visualized by GEPIA2 (p value by one-way ANOVA). Boxplot center represents median, bounds represent 25 and 75%, and whiskers show the minimum or maximum no further than 1.5 * interquartile range from the bound. b, c TRMT6 and TRMT61A protein expression by Western blots in BLCA tumor versus normal samples, with ß-actin as loading control. Relative protein expression is normalized to ß-actin; p values based on two-tailed unpaired student’s t test. d, e m1A mismatch measured by TGIRT-seq in BLCA tumor samples and the paired normal (n = 5). d Overall tRF-3b normalized mismatch index (y-axis, index defined by mismatch% over non-mismatch%) is plot against TRMT6 expression (x-axis, log2 scale from RNA-seq). Significantly higher tRF-3b mismatch index is observed in tumors (p value by paired student’s t test). e tRF-3b A4-mismatch% is compared between tumor and paired normal for each patient (red color in heatmap suggests higher mismatch% in tumor, p value by unpaired two-sided Wilcoxon test). f, g Global de-repression of tRF-3b targets observed in BLCA tumor compared to the paired normal (f, n = 5) or in high-TRMT6 expressed (upper quartile) BLCA tumor (n = 102) compared to the low-TRMT6 expressed (bottom quartile) tumors (n = 102) in TCGA (g). CDF (Cumulative Distribution Function) p value is calculated by one-sided Kolmogorov–Smirnov test to compare overall distribution between tRF-3 targets versus non-targets. h Correlation between TRMT6/61A expression and tRF-3 targets in TCGA BLCA patients (n = 404). Pearson correlation was based on mRNA expression levels in TPM (transcripts per million) by GEPIA2 (visualized on log2 scale). Correlation coefficient summarized by color (red = positive correlation, blue = negative correlation), p value from paired correlation test, *p < 0.05, **p < 0.01. Exact p values from up to down: e 0.0075, 0.0075, 0.0075, 0.025, 0.12, 0.66, 0.66, 0.66, 0.0075, 0.65, 0.66, 0.12, and 0.66; h TRMT6 – 9.1e-8, 7.8e-9, 0.019, 0.78, 8,4e-9, 1.2e-6, 0, 0, 0.005, 4.5e-8; TRMT61A – 9.1e-8, 0.81, 0.08, 0.37, 0.68, 0.94, 0.29, 0.96, 0.17, and 0.47. See also Supplementary Figs. 6, 7 and Supplementary Table 6. Source data are provided as a Source Data file.