title: DNMT3B overexpression by deregulation of FOXO3a-mediated transcription repression and MDM2 overexpression in lung cancer.
abstract: BACKGROUND
  DNA methyltransferase 3B (DNMT3B) contributes to de novo DNA methylation and its overexpression promotes tumorigenesis. However, whether DNMT3B is upregulated by transcriptional deregulation remains unclear.
  
  
  METHODS
  We studied the transcriptional repression of DNMT3B by forkhead O transcription factor 3a (FOXO3a) in lung cancer cell, animal, and clinical models.
  
  
  RESULTS
  The results of luciferase reporter assay showed that FOXO3a negatively regulated DNMT3B promoter activity by preferentially interacting with the binding element FOXO3a-E (+166 to +173) of DNMT3B promoter. Ectopically overexpressed FOXO3a or combined treatment with doxorubicin to induce FOXO3a nuclear accumulation further bound at the distal site, FOXO3a-P (-249 to -242) by chromatin-immunoprecipitation assay. Knockdown of FOXO3a resulted in an open chromatin structure and high DNMT3B mRNA and protein expression. Abundant FOXO3a repressed DNMT3B promoter by establishing a repressed chromatin structure. Note that FOXO3a is a degradation substrate of MDM2 E3-ligase. Cotreatment with doxorubicin and MDM2 inhibitor, Nutlin-3, further enforced abundant nuclear accumulation of FOXO3a resulting in decrease expression of DNMT3B leading to synergistic inhibition of tumor growth and decrease of methylation status on tumor suppressor genes in xenograft specimens. Clinically, lung cancer patients with DNMT3B high, FOXO3a low, and MDM2 high expression profile correlated with poor prognosis examined by immunohistochemistry and Kaplan-Meier survival analysis.
  
  
  CONCLUSIONS
  We reveal a new mechanism that FOXO3a transcriptionally represses DNMT3B expression and this regulation can be attenuated by MDM2 overexpression in human lung cancer model. Cotreatment with doxorubicin and Nutlin-3 is a novel therapeutic strategy through epigenetic modulation.
authors.affiliation: *Department of Pharmacology, College of Medicine, National Cheng Kung University, Tainan, Taiwan; †Institute of Basic Medical Science, College of Medicine, National Cheng Kung University, Tainan, Taiwan; ‡Division of Chest Medicine, Department of Internal Medicine, Chi Mei Medical Center, Tainan; The Center of General Education, Chia Nan University of Pharmacy & Science, Tainan, Taiwan; §Graduate Institute of Pharmacognosy, Taipei Medical University, Taipei, Taiwan; and ‖Division of Thoracic Surgery, Taipei Veterans General Hospital; Institute of Emergency and Critical Care Medicine, National Yang-Ming University School of Medicine, Taipei, Taiwan.
authors.forename: Yi-Chieh
authors.initials: YC
authors.lastname: Yang
authors.forename: Yen-An
authors.initials: YA
authors.lastname: Tang
authors.forename: Jiunn-Min
authors.initials: JM
authors.lastname: Shieh
authors.forename: Ruo-Kai
authors.initials: RK
authors.lastname: Lin
authors.forename: Han-Shui
authors.initials: HS
authors.lastname: Hsu
authors.forename: Yi-Ching
authors.initials: YC
authors.lastname: Wang
mesh_terms: D000818:Animals; D015153:Blotting, Western; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000096:biosynthesis / Q000235:genetics*; D019175:DNA Methylation; D005260:Female; D005500:Follow-Up Studies; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors / Q000096:biosynthesis / Q000235:genetics*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007150:Immunohistochemistry; D008175:Lung Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D018613:Microscopy, Confocal; D009374:Neoplasms, Experimental; D011401:Promoter Regions, Genetic; D051736:Proto-Oncogene Proteins c-mdm2 / Q000096:biosynthesis / Q000235:genetics*; D012334:RNA, Neoplasm / Q000235:genetics*; D012189:Retrospective Studies; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D000097584:DNA Methyltransferase 3B
chemical_list: D004248:DNA (Cytosine-5-)-Methyltransferases; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; D051736:Proto-Oncogene Proteins c-mdm2; D012334:RNA, Neoplasm; D000097584:DNA Methyltransferase 3B; C499517:FOXO3 protein, human; C497206:MDM2 protein, human