Fig 2: Constructing and validating the effects of TRIM6 knockdown and overexpression on cell proliferation in glioma cell lines. (A) Relative mRNA expression levels of TRIM6 in U251 and U373 cells. (B,D) qPCR and Western blot validation of TRIM6 knockdown effect in U251 cell line. (C,E) qPCR and Western blot validation of TRIM6 overexpression effect in U373 cell line. (F,G) The CCK-8 assay was used to detect the cell viability after knocking down TRIM6 in the U251 cell line and overexpressing TRIM6 in the U373 cell line at 24h, 48h, 72h, and 96h. The data represents the mean ± standard deviation (SD) of three independent experiments. (H,I) The colony formation was evaluated using the plate cloning assay after knocking down TRIM6 in the U251 cell line and overexpressing TRIM6 in the U373 cell line. Data are presented as mean ± SD. Statistical analysis was performed using t-test, and p < 0.05 was considered statistically significant (*** p < 0.001).

Fig 3: Investigate the effects of knocking down or overexpressing TRIM6 on cell invasion and migration abilities in glioma cell lines. Graphical representation of wound healing assay results is shown in (A,C). (B,D) The stacked bar graph demonstrates the relative wound width in U251 cells and U373 cells in various treatment groups, respectively. Graphical representation of the Transwell assay results is shown in (E,H). The upper panel illustrates the migration of cells across the Transwell membrane, while the lower panel depicts the invasion of cells through the Matrigel-coated membrane. (F,I) Invasion: The stacked bar graph demonstrates the relative number of invasive U251 cells in various treatment groups. (G,J) Migration: The bar graph represents the relative number of migrated U373 cells in different experimental conditions. Data are presented as mean ± SD. Statistical analysis was performed using t-test, and p < 0.05 was considered statistically significant (*** p < 0.001, **** p < 0.0001).

Fig 4: PPI network analysis and correlation analysis between TRIM6 and hub genes using data from TCGA glioma database. (A) Visual map of the protein-protein interaction network for high and low expression TRIM6 groups. (B) Cytoscape’s plug-in cytoHubba uses the Degree algorithm to select the hub genes from the PPI network. (C) Heatmap demonstrates the correlation between TRIM6 and hub genes. (D) Detection of hub genes expression levels in Glioma tumor tissues and corresponding adjacent non-tumor tissues using qPCR experiment. Data are presented as mean ± SD. Statistical analysis was performed using t−test, and p < 0.05 was considered statistically significant (*** p < 0.001).

Fig 5: Expression analysis and validation of TRIM6. (A–C) Differences in TRIM6 expression between normal and Gliomas, GBM and LGG tissues in TCGA database. (D–E) showed that the transcription level of TRIM6 in Gliomas compared with in normal adjacent tissues from GSE109569 and GSE76070 datasets. (F,G) Validation of the expression level of TRIM6 between normal cerebral cortex and Gliomas tissues using four cases of glioma specimens collected in our center and the Human Protein Atlas database (immunohistochemistry). Data are presented as mean ± deviation (SD). Statistical analysis was performed using t-test, and p < 0.05 was considered statistically significant (* p < 0.05, *** p < 0.001). box-and-whisker plot displays the median (middle line within the box), quartiles (box boundaries), and potential outliers (individual data points).