title: 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.
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.|Department of Otolaryngology-Head and Neck Surgery, The Second People's Hospital of Guangdong Province, Southern Medical University, Guangzhou 510515, China.
authors.forename: Hong
authors.initials: H
authors.lastname: Peng
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.
authors.forename: Yuxiang
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.|Department of Otolaryngology-Head and Neck Surgery, The Second People's Hospital of Guangdong Province, Southern Medical University, Guangzhou 510515, China.
authors.forename: Pinggui
authors.initials: P
authors.lastname: Gong
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.
authors.forename: Longmei
authors.initials: L
authors.lastname: Cai
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.
authors.forename: Xiaoming
authors.initials: X
authors.lastname: Lyu
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Jiang
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.
authors.forename: Jianguo
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.
authors.forename: Juan
authors.initials: J
authors.lastname: Lu
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.
authors.forename: Kaitai
authors.initials: K
authors.lastname: Yao
authors.affiliation: Department of Pathology, The Sixth Affiliated Hospital of Guangzhou Medical University, Qingyuan 511518, China.
authors.forename: Kunping
authors.initials: K
authors.lastname: Liu
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.|Department of Pathology, The Sixth Affiliated Hospital of Guangzhou Medical University, Qingyuan 511518, China.
authors.forename: Jinbang
authors.initials: J
authors.lastname: Li
authors.affiliation: Department of Otorhinolaryngology at Nanfang Hospital, Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Li
mesh_terms: D000368:Aged; D002277:Carcinoma / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D018899:CpG Islands; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000378:metabolism*; D019175:DNA Methylation; D005260:Female; D020868:Gene Silencing; D057809:HEK293 Cells; D004854:Herpesvirus 4, Human / Q000235:genetics; D006801:Humans; D008297:Male; D008875:Middle Aged; D016328:NF-kappa B / Q000378:metabolism*; D000077274:Nasopharyngeal Carcinoma; D009303:Nasopharyngeal Neoplasms / Q000378:metabolism; D051059:PTEN Phosphohydrolase / Q000378:metabolism*; D011485:Protein Binding; D015398:Signal Transduction; D013696:Temperature; D015854:Up-Regulation; D014763:Viral Matrix Proteins / Q000378:metabolism*; D000097584:DNA Methyltransferase 3B
chemical_list: D004248:DNA (Cytosine-5-)-Methyltransferases; D016328:NF-kappa B; D051059:PTEN Phosphohydrolase; D014763:Viral Matrix Proteins; D000097584:DNA Methyltransferase 3B; C037662:EBV-associated membrane antigen, Epstein-Barr virus; C494929:PTEN protein, human
keywords: DNA methylation; DNMT3b; EBV; LMP1; PTEN