TET1 Exerts Its Anti-tumor Functions Via Demethylation DACT2 And SFRP2 To Antagonize Wnt/?-catenin Signaling Pathway In Nasopharyngeal Carcinoma Cells

Objectives The worldwide incidence and mortality of nasopharyngeal carcinoma(NPC)is very low,but it is high in southern China.Radiation therapy is currently the primary treatment in the earlier stage,and combined with chemoradiotherapy in the late stage,but distant metastasis and recurrence are frequent.TET1 is a tumor suppressor gene(TSG)that codes for ten-eleven translocation methyl cytosine dioxygenase1(TET1)catalyzing the conversion of 5-methylcytosine to 5-hydroxy methyl cytosine as a first step of TSG demethylation.Its hypermethylation has been associated with colorectal cancer,renal cancer,gastric cancer and other cancers pathogenesis.However,whether TET1 plays the same role in NPC remains unclear.So we need to explore the mechanism of TET1 role in NPC to provide clinical diagnosis marker of nasopharyngeal carcinoma.Methods Expression and cellular localization of TET1 and 5-hmC were assayed by HE and Immunohistochemical staining.RNA expression and promoter methylation of TET1 in NPC,normal nasal tissues and nasopharyngeal carcinoma cells treated with demethylation reagent Aza with or without TSA histone deacetylase were detected by RT-PCR,qRT-PCR and MSP.MTS assays,colony formation,flow cytometric analysis and trans-well were used to assess various functions(proliferation,induce cycle arrest and apoptosis,migration and invasion)of TET1 in nasopharyngeal carcinoma cells with TET1-CD and TET1-CD-mut transfected.qRT-PCR and immunofluorescence to detect the effect of TET1 on EMT.Western blot,Luciferase Reporter Assay,and immunofluorescence staining assays were used to examine the effect of TET1 on Wnt/?-catenin signaling pathway in nasopharyngeal carcinoma;Dot-blot,qRT-PCR,and MSP assays were used to detect the target gene of TET1 regulation of Wnt/?-catenin signaling pathways.Results TET1 expression was downregulated and hyper-methylation in nasopharyngeal carcinoma(NPC)tissues compared with surgical margin tissues and normal nasal tissues,Demethylation of TET1 in HONE1 and HNE1 cells restored its expression with downregulated methylation,implying that TET1 was silenced by promoter hypermethylation.Ectopic expression of TET1 suppressed the growth of NPC cells,induced apoptosis,arrested cell division in G0/G1 phase,and inhibited cell invasion and migration through regulate EMT,confirming TET1 TSG activity.TET1 decreased the expression of nuclear ?-catenin and downstream target genes.Furthermore,TET1 could cause Wnt antagonists(DACT2,SFRP2)promoter demethylation and restore its expression in NPC cells.Conclusions Collectively,we conclude that TET1 exerts its anti-tumor functions in NPC cells by suppressing Wnt/?-catenin signaling via demethylation of Wnt antagonists(DACT2 and SFRP2).Therefore,TET1 may be used as a biomarker for nasopharyngeal carcinoma,which may contribute to the clinical diagnosis of NPC and the search for other molecular markers.