Fig. 4: tRF-22 binds to and negatively regulates MXD1 expression.

A Volcano plot showed differentially expressed genes following tRF-22 overexpression in U-251 MG cells. (|log2FC| > 2, P < 0.05) 102 genes upregulated (red) and 43 downregulated (green). B Hierarchical clustering heatmap of top 20 downregulated expressed mRNAs regulated by tRF-22 mimic. C The mRNA expression levels of MXD1 in non-tumor and GBM samples were analyzed using the TCGA database. Values are presented as mean ± SD, *P < 0.05 compared to non-tumor samples. D The effect of MXD1 expression levels on the survival of GBM patients was analyzed using the TCGA database. E The differentially expressed genes following tRF-22 overexpression were subjected to GO and KEGG enrichment, sorted by −log10(P). The expression levels of MXD1 in SVG p12, U-87 MG, U-251 MG, and T98G cells were detected by RT-qPCR (F) and western blot (G), respectively. Values are presented as mean ± SD (n = 3), **P < 0.01, ***P < 0.001 compared to the SVG p12 group. H The luciferase activity in 293T cells transfected with wild-type MXD1 3’UTR and tRF-22 binding site mutant dual-luciferase reporter vectors was analyzed using a dual-luciferase reporter system. Values are presented as mean ± SD (n = 5), **P < 0.01 compared to the mimic NC group. I After transfection with tRF-22 mimic and tRF-22 inhibitor in U-251 MG and T98G cells for 48 h, changes in MXD1 expression were detected by Western blot. Values are presented as mean ± SD (n = 3), **P < 0.01 compared to the NC group.