Fig. 4

RUNX2 knockdown in CHIR99021 treated cells decreases proliferation and clonal expansion potential. A Negative correlation of RUNX2 and L1CAM expression in IDH^mut glioma patients (R = –0.313, p = 0.000648) (source: CGGA, mRNA dataset; only primary IDH^mut gliomas included). Correlation was determined using Pearson’s correlation test. B RUNX2 is significantly overexpressed in IDH^wt compared to IDH^mut gliomas (p = 8.5 × 10^− 11) (source: CGGA). C High RUNX2 expression is associated with significantly impaired survival in patients with IDH^mut gliomas (source: Gliovis/CGGA). D qPCR analysis of RUNX2 expression after siRNA-mediated knockdown in CHIR99021-treated IDH^mut astrocytes shows reduced RUNX2 expression compared to control (n = 2 biological and n = 3 technical replicates per sample). E Cell cycle analysis with EdU flow cytometry after siRNA-mediated knockdown of RUNX2 in CHIR99021-treated cells demonstrates a significant decrease of cells in S phase (p = 0.0002–0.0047), a significant increased proportion of cells in G0/G1 (p = 0.0013–0.0161), and inconsistently enhanced proportions of cells in G2/M (p = 0.0331 – ns), (n = 3 biological and 3 technical replicates each). F RUNX2 knockdown in CHIR99021-treated astrocytes leads to significantly reduced clonal expansion potential (p = 0.0062–0.0182) (n = 3 biological and 3 technical replicates each). G Distribution of up- and downregulated DEGs after RUNX2 knockdown in CHIR99021-treated IDH^mut astrocytes, showing a higher fraction of downregulated genes, including several relevant in IDH^mut gliomas. H Ingenuity Pathway analysis of RUNX2-dependent DEGs highlightssignificant enrichment for proliferation-relevant functions, including cellular development, cell death and survival, and cellular growth and proliferation. I Heatmap of selected genes associated with cell death and survival from the siRUNX2 RNA-seq dataset