Higher methylation intensity induced by EBV LMP1 via NF-κB/DNMT3b signaling contributes to silencing of PTEN gene

Abstract

Phosphatase and tensin homolog (PTEN) is a major tumor suppressor and usually silenced via the deletion, insertion and mutation. We previously discovered its inactivation via aberrant CpG island methylation. Here, we provide further evidence that EBV latent membrane protein 1(LMP1) can induce a higher intensity of DNA methylation at PTEN CpG islands, inactivating PTEN at the cellular and molecular level. Initially, increased methylation intensity of PTEN CpG islands was observed in EBV-infected nasopharyngeal carcinoma (NPC) cells, accompanied by decreased PTEN expression. In NPC tissue samples showing the methylation at PTEN promoter, LMP1 was highly expressed in higher methylation intensity group relative to lower intensity group, and DNA methyltransferase 3b (DNMT3b) expression was positively correlated with LMP1 expression. Moreover, transfection of LMP1 gene into EBV-negative NPC cells demonstrated that LMP1 up-regulated DNMT3b expression, leading to a higher intensity of PTEN CpG island methylation. Mechanistically, computational prediction and luciferase reporter assay identified a functional NF-κB binding site on DNMT3b promoter and the mutated NF-κB binding site abolished LMP1-mediated DNMT3b activation. Chromatin immunoprecipitation displayed that NF-κB p65 subunit constitutively bound to DNMT3b promoter, supporting the activation of DNMT3b by EBV LMP1 via NF-κB signaling. Furthermore, the expression level of DNMT3b was observed to be increased in the nuclei of LMP1-expressing NPC cells, and a NF-κB inhibitor, PDTC, counteracted LMP1-mediated DNMT3b overexpression. Thus, this study first reports that LMP1-mediated NF-κB can up-regulate DNMT3b transcription, thereby leading to relatively higher methylation intensity at PTEN CpG islands, and ultimately silencing major tumor suppressor PTEN.

Epstein–Barr virus: a master epigenetic manipulator

“…Expression of LMP1 has been shown to induce DNMTs resulting in DNA hypermethylation and silencing of the CDH1 and PTEN promoters, TSGs also shown to be DNA methylated in NPC and GC [46,47,80]. To define the epigenetic consequence of EBV epithelial infection, we used a transient infection model based on the observation that EBV is not stably maintained in cultured epithelial cells and requires selection pressure for episomal maintenance.…”

EBV Viral Load in Tumor Cells Did Not Predict Tumor Extensiveness in Nasopharyngeal Cancer

“…4 The viral protein LMP1 was associated with inactivation of several important genes such as tumor suppressor genes within the host cells. 4,14,15 The LMP1 inactivated tumor suppressor genes for instance Phosphatase and tensin homolog (PTEN) gene through the action of DNA methyltransferase 3b (DNMT3b). 15 This DNMT3b resulted in PTEN CpG island methylation.…”

“…4,14,15 The LMP1 inactivated tumor suppressor genes for instance Phosphatase and tensin homolog (PTEN) gene through the action of DNA methyltransferase 3b (DNMT3b). 15 This DNMT3b resulted in PTEN CpG island methylation. 15 Furthermore, the LMP1 expressed by EBV has also been shown to be able to methylate CDH-1 promoter region which eventually silencing E-cadherin gene.…”

“…15 This DNMT3b resulted in PTEN CpG island methylation. 15 Furthermore, the LMP1 expressed by EBV has also been shown to be able to methylate CDH-1 promoter region which eventually silencing E-cadherin gene. The cadherin is cell adhesion molecule in which deficient in this protein will promote cancer cells to easily metastasize.…”

Hypermethylation of brain natriuretic peptide gene is associated with the risk of rheumatic heart disease

“…DNA methylation often occurs in a CpG dinucleotide context and promoter DNA methylation can regulate the expression level of the gene [6,7]. Aberrant DNA methylation has been extensively studied in cancer, lung cancer and leukaemia [8–10].…”