Abstract
BackgroundNasopharyngeal carcinoma (NPC) is prevalent in South China, including Hong Kong and Southeast Asia, constantly associated with Epstein-Barr virus (EBV) infection. Epigenetic etiology attributed to EBV plays a critical role in NPC pathogenesis. Through previous CpG methylome study, we identified Disheveled-associated binding antagonist of beta-catenin 2 (DACT2) as a methylated target in NPC. Although DACT2 was shown to regulate Wnt signaling in some carcinomas, its functions in NPC pathogenesis remain unclear.MethodsRT-PCR, qPCR, MSP, and BGS were applied to measure expression levels and promoter methylation of DACT2 in NPC. Transwell, flow cytometric analysis, colony formation, and BrdU-ELISA assay were used to assess different biological functions affected by DACT2. Immunofluorescence, Western blot, and dual-luciferase reporter assay were used to explore the mechanisms of DACT2 functions. Chemosensitivity assay was used to measure the impact of DACT2 on chemotherapy drugs.ResultsWe found that DACT2 is readily expressed in multiple normal adult tissues including upper respiratory tissues. However, it is frequently downregulated in NPC and correlated with promoter methylation. DNA methyltransferase inhibitor 5-aza-2′-deoxycytidine restored its expression in NPC cells. DACT2 methylation was further detected in 29/32 (91%) NPC tumors but not in any (0/8) normal nasopharyngeal tissue samples. Ectopic expression of DACT2 in NPC cells suppressed their proliferation, migration, and invasion through downregulating matrix metalloproteinases. DACT2 expression also induced G2/M arrest in NPC cells through directly suppressing β-catenin/Cdc25c signaling, which sensitized NPC cells to paclitaxel and 5-FU, but not cisplatin.ConclusionOur results demonstrate that DACT2 is frequently inactivated epigenetically by CpG methylation in NPC, while it inhibits NPC cell proliferation and metastasis via suppressing β-catenin/Cdc25c signaling. Our study suggests that DACT2 promoter methylation is a potential epigenetic biomarker for the detection and chemotherapy guidance of NPC.
Highlights
Nasopharyngeal carcinoma (NPC) is prevalent in South China, including Hong Kong and Southeast Asia, constantly associated with Epstein-Barr virus (EBV) infection
Disheveled-associated binding antagonist of beta-catenin 2 (DACT2) was downregulated in NPC by promoter methylation Reverse transcription (RT)-PCR confirmed that DACT2 was expressed in the majority of normal adult tissues (Fig. 1a)
DACT2 expression was downregulated in HNE1 and HONE1 NPC cells and was restored by 5aza-2′-deoxycytidine (Aza) without or with trichostatin A (TSA)
Summary
Nasopharyngeal carcinoma (NPC) is prevalent in South China, including Hong Kong and Southeast Asia, constantly associated with Epstein-Barr virus (EBV) infection. Through previous CpG methylome study, we identified Disheveled-associated binding antagonist of beta-catenin 2 (DACT2) as a methylated target in NPC. DACT2 was shown to regulate Wnt signaling in some carcinomas, its functions in NPC pathogenesis remain unclear. The Disheveled-associated binding antagonist of β-catenin (DACT) family, known as Dapper/ Frodo, are small intracellular scaffold proteins. Our findings demonstrated that DACT2 antagonizes Akt/GSK-3 and Wnt/β-catenin signaling to suppress epithelial-to-mesenchymal transition (EMT) [3]. DACT2 interacts with Wnt/β-catenin signaling to prevent Yes-associated protein translocation to the nucleus, resulting in its sequestration and degradation in the cytoplasm [8]. Zebrafish DACT2 was reported to inhibit TGF-β/Nodal signaling during mesoderm induction by interacting with type 1 receptors ALK5 and ALK4 and further promoting lysosomal degradation [10, 11]
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