Extended Data Fig. 6: The pseudouridine modification profile in GSCs.
From: Targeting PUS7 suppresses tRNA pseudouridylation and glioblastoma tumorigenesis
(a) A representative PUS7-dependent pseudouridine site identified by small RNA DM-Ψ-seq in PBT003 GSCs. (b) Validation of the PUS7-dependent pseudouridine site in tRNA-Arg-CCG-2-1 in PUS7 KO PBT003 GSCs by primer extension assay. The uncropped blot images for the cropped images shown here are in the source data. Repeated twice with similar results. (c) A representative PUS7-dependent pseudouridine site in tRNA-Glu-TTC-4-1 in control or C17-treated PBT003 GSCs. (d) Pearson correlation analysis for global tRNA abundance in control and PUS7 KO PBT003 GSCs. (e) Expression of tRNA-Arg-CCG in control and PUS7 KO PBT003 GSCs examined by Northern blot analysis. U6 was used as a loading control. The uncropped blot images for the cropped images shown here are in the source data. Repeated twice with similar results. (f) Analysis of tRF abundance in control and PUS7 KO PBT003 GSCs. Red dots: tRFs derived from tRNAs with PUS7-dependent pseudouridine sites. The q value was calculated by Cochran Mantel Haenszel test and adjusted by BH methods. (g) The OP-puro incorporation analysis of control and PUS7 KO PBT707 GSCs. (h) Nascent protein synthesis and total protein level analysis of control and PUS7 KO 293T cells. (i) A luciferase reporter assay to test tRNA translation efficiency in control and PUS7 KO PBT707 cells. n = 3 cell culture replicates. Error bars are SE of the mean. *p < 0.05 (p = 0.0251), and ns: not statistically significant [p > 0.05, p = 0.0921 for control, p = 0.0251 for 6x(CGG)Arg, and p = 0.1238 for 6x(CGA)Arg] by one-tailed Student’s t-test.
