apaf1  (Proteintech)

Journal: BMC Musculoskeletal Disorders

Article Title: The METTL3-IGF2BP3 axis drives osteosarcoma progression by enhancing ID1 mRNA stability

doi: 10.1186/s12891-026-09527-0

Figure Lengend Snippet: Identification of OS-specific RNA-binding proteins regulating ID1 and mechanistic predictions. A : Volcano plot of DEGs in the GSE253548 dataset. B : Venn diagram intersecting ENCORI-predicted RBPs with DEGs. C : Feature selection by LASSO regression. D : SVM-RFE algorithm-derived feature genes. E : Variable importance ranking from random forest analysis. F : Venn diagram overlapping feature genes identified by LASSO, SVM-RFE, and random forest. G : IGF2BP3 expression profile in the GSE253548 dataset. H : RT-qPCR analysis of IGF2BP3 mRNA levels. I : Western blot detection of IGF2BP3 protein expression. J : SRAMP-predicted m 6 A modification site on ID1 mRNA. K : Secondary structure of the m 6 A-modified site at 782 nt on ID1 mRNA. L : MeRIP-qPCR quantification of ID1 m 6 A modification levels. M : RM2Target-predicted m 6 A-mediated interaction between IGF2BP3 and ID1. N : Spearman correlation of IGF2BP3 and ID1 expression. Data are presented as mean ± SEM. ** P < 0.01, *** P < 0.001, **** P < 0.0001. Statistical analyses were performed using independent-samples t -test or one-way ANOVA followed by Tukey’s post hoc test. All experiments were performed in triplicate.

Article Snippet: Following lysis of infected MG-63 cells, supernatants were incubated overnight at 4 °C with pre-blocked Protein A/G agarose beads conjugated to anti-IGF2BP3 antibody (81805-1-RR, Proteintech), anti-METTL3 (#86132, Cell Signaling Technology, Danvers, MA, USA), or control IgG (A7016, Beyotime).

Techniques: RNA Binding Assay, Selection, Derivative Assay, Expressing, Quantitative RT-PCR, Western Blot, Modification

Journal: BMC Musculoskeletal Disorders

Article Title: The METTL3-IGF2BP3 axis drives osteosarcoma progression by enhancing ID1 mRNA stability

doi: 10.1186/s12891-026-09527-0

Figure Lengend Snippet: IGF2BP3 regulates ID1 to modulate malignant behaviors in OS Cells. A - B : RT-qPCR analysis of IGF2BP3 expression in engineered cells. C : RT-qPCR detection of ID1 mRNA levels. D : Western blot analysis of ID1 protein expression. E : RIP-qPCR demonstrating IGF2BP3-ID1 mRNA interaction, with RIP efficiency validated by Western blot. F : Schematic of ID1-WT/MUT CDS and their dual-luciferase reporter constructs. G : Dual-luciferase reporter gene assay results. H : RT-qPCR for ID1 mRNA expression in cells at different time points (0, 2, 4, 6 h) after actinomycin D treatment. I : Western blot for IGF2BP3 and ID1 expression in cells. J : CCK-8 assay assessing cellular proliferation. K : Colony formation capacity quantified across experimental groups. L : Scratch assay evaluating cell migration ability, scale bar = 400 μm. M : Transwell invasion assay with representative images, scale bar = 200 μm. Data are presented as mean ± SEM. ns P > 0.05, **** P < 0.0001. Statistical analyses were performed using independent-samples t-test or one-way ANOVA followed by Tukey’s post hoc test. All experiments were performed in triplicate.

Article Snippet: Following lysis of infected MG-63 cells, supernatants were incubated overnight at 4 °C with pre-blocked Protein A/G agarose beads conjugated to anti-IGF2BP3 antibody (81805-1-RR, Proteintech), anti-METTL3 (#86132, Cell Signaling Technology, Danvers, MA, USA), or control IgG (A7016, Beyotime).

Techniques: Quantitative RT-PCR, Expressing, Western Blot, Luciferase, Construct, Reporter Gene Assay, CCK-8 Assay, Wound Healing Assay, Migration, Transwell Invasion Assay

Journal: BMC Musculoskeletal Disorders

Article Title: The METTL3-IGF2BP3 axis drives osteosarcoma progression by enhancing ID1 mRNA stability

doi: 10.1186/s12891-026-09527-0

Figure Lengend Snippet: The METTL3-IGF2BP3 axis mediates m 6 A-dependent regulation of ID1 expression and stability. A : METTL3 expression in GSE253548 dataset. B - C : METTL3 expression in cell lines via RT-qPCR (B) and Western blot (C). D : RM2Target prediction of METTL3-ID1 interaction. E : Spearman correlation between METTL3 and ID1 expression. F - G : METTL3 expression validation in engineered cells. H : MeRIP-qPCR analysis of ID1 m6A modification levels. I : RIP-qPCR analysis of the interaction between METTL3 and ID1 mRNA, with the efficiency of the RIP assay validated by Western blot. J : RIP-qPCR detection of IGF2BP3-ID1 mRNA interaction. K : MeRIP-qPCR analysis of ID1 m6A modification levels in MG-63 cells with endogenous METTL3 knockdown. L : RIP-qPCR analysis of the interaction between IGF2BP3 and ID1 mRNA in MG-63 cells with endogenous METTL3 knockdown. M - N : ID1 mRNA (M) and protein (N) expression under indicated treatments. O : RNA stability assay under actinomycin D treatment (0, 2, 4, 6 h). Data are presented as mean ± SEM. ns P > 0.05, *** P < 0.001, **** P < 0.0001. Statistical analyses were performed using independent-samples t-test or one-way ANOVA followed by Tukey’s post hoc test. All experiments were performed in triplicate.

Article Snippet: Following lysis of infected MG-63 cells, supernatants were incubated overnight at 4 °C with pre-blocked Protein A/G agarose beads conjugated to anti-IGF2BP3 antibody (81805-1-RR, Proteintech), anti-METTL3 (#86132, Cell Signaling Technology, Danvers, MA, USA), or control IgG (A7016, Beyotime).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Biomarker Discovery, Modification, Knockdown, Stability Assay

Journal: BMC Musculoskeletal Disorders

Article Title: The METTL3-IGF2BP3 axis drives osteosarcoma progression by enhancing ID1 mRNA stability

doi: 10.1186/s12891-026-09527-0

Figure Lengend Snippet: The METTL3-IGF2BP3 axis mediates ID1-dependent malignant behaviors in OS cells. A : CCK-8 assay assessing cell proliferation. B : Colony formation assay quantifying clonogenic capacity. C : Scratch wound healing assay evaluating migration, scale bar = 400 μm. D : Transwell invasion assay, scale bar = 200 μm. E : Cell proliferation evaluated by CCK8 assay. F : Colony formation assay to assess colony numbers. G : Scratch assay to evaluate the migration ability of cells, scale bar = 400 μm. H : Transwell assay to assess the invasion ability of cells, scale bar = 200 μm. Data are presented as mean ± SEM. ** P < 0.01, *** P < 0.001, **** P < 0.0001. Statistical analyses were performed using independent-samples t-test or one-way ANOVA followed by Tukey’s post hoc test. All experiments were performed in triplicate.

Article Snippet: Following lysis of infected MG-63 cells, supernatants were incubated overnight at 4 °C with pre-blocked Protein A/G agarose beads conjugated to anti-IGF2BP3 antibody (81805-1-RR, Proteintech), anti-METTL3 (#86132, Cell Signaling Technology, Danvers, MA, USA), or control IgG (A7016, Beyotime).

Techniques: CCK-8 Assay, Colony Assay, Wound Healing Assay, Migration, Transwell Invasion Assay, Transwell Assay

Journal: BMC Musculoskeletal Disorders

Article Title: The METTL3-IGF2BP3 axis drives osteosarcoma progression by enhancing ID1 mRNA stability

doi: 10.1186/s12891-026-09527-0

Figure Lengend Snippet: Effects of METTL3 and IGF2BP3 inhibitors on ID1-mediated malignant behaviors of OS cells. A : RT-qPCR analysis of ID1 expression in each group of cells. B : WB analysis of ID1 expression in each group of cells. C : RT-qPCR analysis of ID1 mRNA levels at different time points (0, 2, 4, 6 h) following actinomycin D treatment. D : CCK-8 assay assessing cell proliferation. E : Colony formation assay evaluating colony numbers in each group. F : Wound healing assay assessing cell migration ability; scale bar = 400 μm. G : Transwell assay assessing cell invasion ability; scale bar = 200 μm. Data are presented as mean ± SEM. **** P < 0.0001. Statistical analyses were performed using one-way ANOVA followed by Tukey’s post hoc test. All cell experiments were repeated three times.

Article Snippet: Following lysis of infected MG-63 cells, supernatants were incubated overnight at 4 °C with pre-blocked Protein A/G agarose beads conjugated to anti-IGF2BP3 antibody (81805-1-RR, Proteintech), anti-METTL3 (#86132, Cell Signaling Technology, Danvers, MA, USA), or control IgG (A7016, Beyotime).

Techniques: Quantitative RT-PCR, Expressing, CCK-8 Assay, Colony Assay, Wound Healing Assay, Migration, Transwell Assay

Journal: BMC Musculoskeletal Disorders

Article Title: The METTL3-IGF2BP3 axis drives osteosarcoma progression by enhancing ID1 mRNA stability

doi: 10.1186/s12891-026-09527-0

Figure Lengend Snippet: The METTL3-IGF2BP3 axis mediates ID1-dependent tumor growth in vivo. A : Western blot analysis of METTL3 and ID1 expression in tumor tissues from each group of mice. B : Western blot analysis of IGF2BP3 and ID1 expression in tumor tissues from each group of mice. C - D : Changes in tumor volume in each group of mice. E - F : Representative images of tumors from each group of mice. G - H : Tumor weights in each group of mice. I - J : Representative immunohistochemical images of Ki67 staining and quantification of Ki67-positive area percentage in tumor tissues from each group of mice, scale bar = 100 μm. N = 6. Data are presented as mean ± SEM. ns P > 0.05, *** P < 0.001, **** P < 0.0001. Statistical analyses were performed using one-way ANOVA followed by Tukey’s post hoc test.

Article Snippet: Following lysis of infected MG-63 cells, supernatants were incubated overnight at 4 °C with pre-blocked Protein A/G agarose beads conjugated to anti-IGF2BP3 antibody (81805-1-RR, Proteintech), anti-METTL3 (#86132, Cell Signaling Technology, Danvers, MA, USA), or control IgG (A7016, Beyotime).

Techniques: In Vivo, Western Blot, Expressing, Immunohistochemical staining, Staining

Journal: BMC Musculoskeletal Disorders

Article Title: The METTL3-IGF2BP3 axis drives osteosarcoma progression by enhancing ID1 mRNA stability

doi: 10.1186/s12891-026-09527-0

Figure Lengend Snippet: Mechanistic diagram of METTL3-IGF2BP3-mediated ID1 in promoting osteosarcoma progression

Article Snippet: Following lysis of infected MG-63 cells, supernatants were incubated overnight at 4 °C with pre-blocked Protein A/G agarose beads conjugated to anti-IGF2BP3 antibody (81805-1-RR, Proteintech), anti-METTL3 (#86132, Cell Signaling Technology, Danvers, MA, USA), or control IgG (A7016, Beyotime).

Techniques: