Supplementary Figure 6: Leukemia oncogenes induce SnoRNA expression.
From: AML1-ETO requires enhanced C/D box snoRNA/RNP formation to induce self-renewal and leukaemia
(a) RT-PCR analysis shows expression of snoRNAs in AE9a transduced lin− bone marrow cells, compared to control. (b) and (c) Scatterplot shows snoRNA expression in myc- (b), MLL/AF9-transduced (c) and control (empty vector) lin− bone marrow cells determined by snoRNA-seq. Paired Wilcoxon test. (d) RT-PCR analysis of Aes mRNA expression in lin− bone marrow cells transduce with AE9a, MLL-AF9 or c-Myc, compared to control. (e) and (f) Scatterplot shows snoRNA expression in MLL-AF9 (e) and c-Myc (f) transduced AES wildtype (AES WT) and knockout (AES KO) lin− bone marrow cells determined by snoRNA-seq. Lin− bone marrow cells isolated from ROSA26-Cre/ERT; Aes+/+ or ROSA26-Cre/ERT; Aesf/f were retrovirally transduced with oncogene. Transduced cells were treated with 4-hydroxytamoxifen for three days for Aes deletion. (g) Kaplan–Meier survival analysis of mice injected with MLL-AF9 transduced lin− bone marrow cells isolated from Cre/ERT; Aes+/+ or ROSA26-Cre/ERT; Aesf/f mice. Tamoxifen induction was performed at two weeks after transplantation. N = 7 and 8 mice, log-rank test. (h) Kaplan–Meier survival analysis of mice injected with c-Myc transduced lin− bone marrow cells isolated from AES wildtype (Aes+/+) or straight knockout (Aes−/−) mice. N = 12 mice each group, log-rank test. (i) Cell cycle analysis of U937 cells expressing empty vector (MIG) or AML1-ETO. One of n = 3 independent experiment is shown. (j) Cell cycle distribution of control (MIG) or AML1-ETO-expressing (AE) U937 cells. (k) Scatterplot shows snoRNA expression in control (MIG) or AML-ETO-expressing U937 cells determined by snoRNA-seq. (l) RT-PCR analysis shows expression of snoRNAs in control (MIG) or AML-ETO-expressing (AE) U937 cells. (m) Pseudouridylation ratios were analyzed at 72 sites in 18S and 28S rRNA in human primary AML1-ETO (n = 5) and CD34+ cell samples (n = 5). (n) Heatmap shows significant snoRNAs associated with chemotherapy response. Error bars, mean ± s.d. of n = 3 independent experiments unless otherwise indicated. Statistical source data for Supplementary Fig. 6j are provided in Supplementary Table 7.
