Fig. 4
Isolating oncogenic targets of miR-212-3p in group 3 MB tumors. A Venn diagram showing 37 common targets between miRNA target prediction databases (Targetscan and miRDip) and genes downregulated (Log2 fold change < − 1.0, p < 0.05) in miR-212 expressing HDMB03 cells (via RNA sequencing analysis). B Expression heat map of the 37 identified targets of miR-212 in two MB data sets, (i) local (pediatric cerebellum n = 10; group 3 MB n = 7; Kanchan et al., GSE148390) and (ii) larger meta-dataset (normal cerebellum n = 291; group 3 MB n = 233; Weishaput et al., GSE124814). C Elevated expression of NFIB across MB datasets (NC n = 9, Roth et al. 2008; MB1 n = 76, Gilbertson et al. 2012; MB2 n = 223, Pfister et al. 2017; MB3 n = 57, Delattre et al. 2012; MB4 n = 62, Kool et al. 2009). D Elevated NFIB expression across subgroups within local MB cohort (pediatric NC n = 10; SHH n = 6; group 3 n = 7; group 4 n = 12; Kanchan et al., GSE148390). E Kaplan–Meier Survival Analysis demonstrating poor survival in high NFIB-expressing MB patients (n = 542) compared to low-expressing patients (n = 70) (Cavalli et al., GSE85217). F Western blotting showing significantly increased expression of NFIB in classic group 3 MB cell lines (HDMB03, D425, and D341). G Immunohistochemical staining of NFIB showing intense nuclear expression in group 3 MB patient samples (n = 9) when compared to normal cerebellum (n = 8). H Dual-luciferase assay confirming NFIB as a direct target of miR-212-3p. I RT-PCR and J Western blotting analysis showing significantly decreased expression of NFIB in miR-212-3p transiently transfected HDMB03 cells. β-actin served as an internal loading control. Data presented as mean ± SD from experiments done in triplicate and analyzed using Mann–Whitney U test (C and D) or Student’s t-test (E, G, and H); *p < 0.05, **p < 0.01, ***p < 0.001. Scale bar: 100 μm. NC, normal cerebellum