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. 2025 Jul 28;21(11):4942–4960. doi: 10.7150/ijbs.112133

Figure 4.

Figure 4

IGF2BP3 interacts with and stabilizes MALAT1 in a m6A-dependent manner. A Venn diagram showing the overlap of DEGs from GSE6764, GSE94660, TCGA-LIHC cohort, and starBase database. B Relative expression of the six candidate RBPs was detected by qRT-PCR in HepG2.2.15 and HBx-expressing HepG2 cells. C Correlation analysis showing a positive relationship between IGF2BP3 and MALAT1 in the TCGA-LIHC cohort. D Binding ability of MALAT1 and IGF2BP3 was predicted by RPISeq. E Interaction between MALAT1 and IGF2BP3 was predicted by HDOCK. F Representative images showing the colocalization of MALAT1 (red) and IGF2BP3 (green) in HepG2.2.15 and HBx-expressing HepG2 cells. Scale bars: 20 μm. G RNA pulldown assays followed by silver staining of protein extracts from HBx-expressing HepG2 cells. H RNA pulldown assays were performed in HepG2.2.15 and HepG2 cells. I RIP assays were performed in HepG2.2.15 and HepG2 cells. J RIP assays were performed in HepG2.2.15 cells transfected with Cas9-METTL3, Cas9-METTL14, and Cas9-WTAP plasmids. K Dual-luciferase reporter assays were used to confirm the interaction between IGF2BP3 and two mutants (6021C and 7265C) of the MALAT1 m6A. L RIP assays were performed in HepG2.2.15 cells transfected with MALAT1 containing the indicated mutations. M Scheme of Flag-tagged full-length IGF2BP3 (F#6) and the five truncated mutants (F#1: 1-77aa; F#2: 1-156aa; F#3: 1-262aa; F#4:1-344aa; F#5:1-471aa) were constructed (left). In vitro binding assays showing the enriched MALAT1 in HepG2.2.15 cells detected by RT-PCR (right, upper panel) after incubation with full-length or truncated Flag-tagged IGF2BP3 protein validated by WB (right, lower panel). N RIP assays were performed in HepG2.2.15 cells transfected with plasmids containing the full-length or truncated constructs. O RNA pulldown assays were performed in HepG2.2.15 cells transfected with plasmids containing the indicated full-length or truncated constructs. P The RNA half-life of MALAT1 was measured by performing RNA decay assaysin HepG2.2.15 and HBx-expressing HepG2 cells with IGF2BP3 overexpression or knockdown. *P < 0.05; **P < 0.01; ***P < 0.001. n.s., not significant.