Fig 2: Methyltransferase‐like 14 (METTL14) suppresses renal cell carcinoma tumor growth and metastasis in vivo. A, Subcutaneous xenograft models were established with METTL14 overexpressing (OE‐METTL14) or control (OE‐NC) 769‐P cells (n = 6). B, C, Growth curve (B) and weight (C) of OE‐METTL14 tumors compared to control group (OE‐NC). D, Immunohistochemical assays showing METTL14 and Ki‐67 expression in each group. Scale bar, 200 μm. E, F, Representative metastatic lung nodules in each group and quantification of the number of metastatic nodules. Magnification, 400×. Error bars represent mean ± SD. Mann‐Whitney U test (C, F) and two‐way ANOVA test (B) were used. *P < .05, **P < .01, ***P < .001

Fig 3: Methyltransferase‐like 14 (METTL14) negatively regulates nuclear enriched abundant transcript 1_1 (NEAT1_1) in an N6‐methyladenosine (m6A)‐dependent manner. A, m6A levels of total RNAs in tumor and matched normal tissues by RNA m6A colorimetry (n = 10). B, m6A levels of total RNAs in renal cell carcinoma (RCC) cell lines and HK‐2 cells by colorimetry. C, Correlation analysis between m6A and METTL14 in RCC samples (n = 20). D, m6A levels of METTL14 knockdown or overexpressing (OE) RCC cells by m6A dot blot assay. MB, methylene blue; NC, control. E, Methylated‐RNA immunoprecipitation (IP) assay‐quantitative PCR analysis of NEAT1_1 in METTL14 overexpressing (OE‐METTL14) RCC cells and the control (OE‐NC). F, NEAT1_1 expression in RCC cells treated with 3‐deazaadenosine (DAA) and the control. Error bars represent the mean ± SD. Student’s t test was used. *P < .05, **P < .01, ***P < .001

Fig 4: Low expression of methyltransferase‐like 14 (METTL14) is related to renal cell carcinoma (RCC) progression and poor prognosis. A, B, METTL14 mRNA (n = 47) and protein (n = 6) expression in RCC (T) and matched normal (N) tissues. C, METTL14 expression in KIRC from The Cancer Genome Atlas (TCGA) database (n = 533). D, E, Immunohistochemical (IHC) analysis of METTL14 in RCC tissue microarrays (n = 132). Magnification, 100×, 400×. F‐H, METTL14 expression in tumors >4 cm group (n = 100) vs tumors ≤4 cm group (n = 110), in the RCC with metastasis group (n = 50) vs RCC without metastasis group (n = 160), and in the Fuhrman I/II group (n = 144) vs Fuhrman III/IV group (n = 66). I, J, Kaplan‐Meier analysis of correlation between METTL14 and overall survival (OS) or recurrence‐free survival. K, Multivariate analysis of factors associated with OS in RCC patients (n = 210). L, Time‐dependent OS receiver operating characteristic (ROC) analysis. Error bars represent mean ± SD. Student’s t test (A, E‐H) and log‐rank test (I, J) were used. *P < .05, **P < .01, ***P < .001. AUC, area under the ROC curve; CI, confidence interval; HR, hazard ratio

Fig 5: Methyltransferase‐like 14 (METTL14) inhibits the proliferation and migration of renal cell carcinoma (RCC) cells in vitro. A, B, METTL14 mRNA (A) and protein (B) levels in 769‐P and 786‐O cells transfected with METTL14 siRNAs (si‐M14‐1 and si‐M14‐2) or control (si‐NC). C‐E, CCK‐8 assay (C), clone formation assay (D), and Transwell migration assay (E) of 769‐P and 786‐O cells transfected with METTL14 siRNAs or control. F, G, METTL14 mRNA (F) and protein (G) levels in METTL14 overexpression (OE‐M14) and control (OE‐M14) RCC cells (P < .001). H‐J, CCK‐8 assay (H), cell clone formation assay (I), and Transwell migration assay (J) of METTL14 overexpression and control RCC cells. Error bars represent mean ± SD of at least three independent experiments. Two‐way ANOVA (C, H) and Student’s t test (A, D‐F, I, J) were used. *P < .05, **P < .01, ***P < .001