Fig 2: Clinical significance of risk grouping in TCGA-ESCA database (A) The heatmap shows the clinicopathological features were compared between the high- and low-risk groups. (B) Univariate Cox analysis of the risk score and clinicopathological features. (C) Multivariate Cox analysis of the risk score and clinicopathological features to identify the independent prognostic predictors in TCGA-ESCA database. (D) Kaplan-Meier analysis for patients of TCGA-ESCA database in high- and low-risk group. Data are shown as the means ± SD. *p < 0.05. ESCA, esophageal squamous cell carcinoma; TCGA, The Cancer Genome Atlas; HNRNPC, RNA-binding protein, is a member of the heterogeneous ribonucleoproteins C; YTHDF1, YTH N6-methyladenosine RNA-binding protein 1.

Fig 3: Identify ESCC-related m6A regulators, build risk models, and validate model predictive capabilities (A) Venn diagrams determined YTHDF1 and HNRNPC as ESCC risk factors after overlapping m6A regulators with higher meanDecreaseGini score in both RNA-seq data and TCGA-ESCA database. (B) Nomogram was constructed using YTHDF1 and HNRNPC to visualize the predictive ability of ESCC-related m6A regulators for Normal group and ESCC group. (C) Calibration curve with nomogram predicted probabilities on the x-axis and actual probabilities on the y-axis. The dashed line represents the true value, and the solid line is bias-corrected by Bootstrapping (1000 repetitions). (D) ROC curve was applied to assess the predictive efficiency of the model. (E) Clinical decision curve. The red line represents the outcome predicted by the model, the grey line represents “all positive diagnosis (ESCC)” and the black line represents “all negative diagnosis (Normal)”. The horizontal axis is Threshold Probability, and the vertical axis is the net profit rate after subtracting the disadvantages. (F) Clinical Impact Curve (CIC). The red curve (Number of high risk) represents the number of people classified as ESCC by the model at each threshold probability; the blue curve (Number high risk with outcome) represents the number of true ESCC at each threshold probability. AUC, Area Under Curve; CIC, Clinical Impact Curves; ESCC/ESCA, Esophageal quamous cell carcinoma; HNRNPC, RNA-binding protein, is a member of the heterogeneous ribonucleoproteins C; m6A, N6-methyladenosine; TCGA, The Cancer Genome Atlas; YTHDF1, YTH N6-methyladenosine RNA-binding protein 1.

Fig 4: HNRNPC knockdown attenuated the cell clonogenicity and proliferation in vitro. a We established stable HNRNPC-knockdown cell lines (A549, H1299, and PC-9) by transfecting cells with the lentivirus. WB analysis confirmed that the expression of HNRNPC was inhibited after HNRNPC knockdown (n = 3). b, c clone formation assay revealed that HNRNPC knockdown attenuated the cell clonogenicity of NSCLC after 2 weeks (n = 3). d MTS assay showed that HNRNPC knockdown suppressed the proliferation of NSCLC cells after 24 h (n = 3). *P < 0.05

Fig 5: HNRNPC was involved in cell growth, cell migration, extracellular matrix organization and angiogenesis. a We constructed H1299 cells with stably knockdown of HNRNPC (shHNRNPC) and control cells (sh-NC). By RNA-seq analysis, 10,907 differentially expressed genes were identified, including 5653 upregulated and 5254 downregulated genes. b The GO analysis revealed that the differentially expressed genes were enriched in cell migration, angiogenesis, extracellular matrix organization, cell proliferation, T cell mediated cytotoxicity and collagen fibril organization. c–h The GSEA analysis demonstrated that the cell growth, cell migration, extracellular matrix organization, angiogenesis, collagen fibril organization and T cell mediated cytotoxicity were correlated with HNRNPC knockdown