J Cancer 2022; 13(6):1972-1984. doi:10.7150/jca.66830 This issue
1. Research Center, the Fourth Affiliated Hospital of Hebei Medical University, Shijiazhuang, Hebei, 050011, People's Republic of China
2. Department of Thoracic Surgery, the Fourth Affiliated Hospital of Hebei Medical University, Shijiazhuang, Hebei, 050011, People's Republic of China
Background: Esophageal squamous cell carcinoma (ESCC) is one of the most common aggressive tumors in the world. m6A modification has been implicated to play an important role in many biological progressions. METTL3 as the main methyltransferase has been found in many cancers, including ESCC. Here, we investigated the underlying mechanism of METTL3 in the development of ESCC.
Methods: Quantitative real-time PCR (qRT-PCR), immunohistochemical (IHC) and western blot were used to detect METTL3 expression. To evaluate the function of METTL3, MTS, colony formation, scratch wound healing assay, and transwell and invasion assays were performed. To find out the downstream target of METTL3, mRNA sequencing (mRNA-seq) was conducted. GO and KEGG functional enrichment analyses were carried out to predict possible biological processes and signaling pathways. qRT-PCR and western blot were performed to identify the expression of COL12A1 and the phosphorylation status of RAF, MRK and ERK. Cotransfection of small interfering RNA (for METTL3 silence) with plasmid (for overexpression of COL12A1) and the following gain- and loss-of-function experiments were performed to detect the target gene function of COL12A1 in progression of ESCC mediated by METTL3.
Results: Using TCGA database, higher METTL3 expression was found in ESCC tissues. Moreover, we found that METTL3 was significantly increased in ESCC patient tissues compared with normal tissues and correlated with poor prognosis. The expression of METTL3 in ESCC cell lines was assessed. The gain- and loss-of-function indicates that METTL3 promotes cell proliferation, migration and invasion. Additionally, we confirmed that METTL3 can promote the expression of COL12A1 and upregulate the phosphorylation of RAF, MER and ERK, and moreover COL12A1 can restrain siMETTL3-mediated inhibition of proliferation, migration and invasion in ESCC.
Conclusion: Our study revealed that METTL3 may have an oncogenic role, facilitating the ESCC progression and metastasis by COL12A1/MAPK signaling pathway.
Keywords: esophageal squamous cell carcinoma (ESCC), METTL3, COL12A1, MAPK, proliferation, invasion