title: High-risk human papillomavirus E6 protein promotes reprogramming of Fanconi anemia patient cells through repression of p53 but does not allow for sustained growth of induced pluripotent stem cells.
abstract: DNA repair plays a crucial role in embryonic and somatic stem cell biology and cell reprogramming. The Fanconi anemia (FA) pathway, which promotes error-free repair of DNA double-strand breaks, is required for somatic cell reprogramming to induced pluripotent stem cells (iPSC). Thus, cells from Fanconi anemia patients, which lack this critical pathway, fail to be reprogrammed to iPSC under standard conditions unless the defective FA gene is complemented. In this study, we utilized the oncogenes of high-risk human papillomavirus 16 (HPV16) to overcome the resistance of FA patient cells to reprogramming. We found that E6, but not E7, recovers FA iPSC colony formation and, furthermore, that p53 inhibition is necessary and sufficient for this activity. The iPSC colonies resulting from each of these approaches stained positive for alkaline phosphatase, NANOG, and Tra-1-60, indicating that they were fully reprogrammed into pluripotent cells. However, FA iPSC were incapable of outgrowth into stable iPSC lines regardless of p53 suppression, whereas their FA-complemented counterparts grew efficiently. Thus, we conclude that the FA pathway is required for the growth of iPSC beyond reprogramming and that p53-independent mechanisms are involved.
  
  
  OBJECTIVE
  A novel approach is described whereby HPV oncogenes are used as tools to uncover DNA repair-related molecular mechanisms affecting somatic cell reprogramming. The findings indicate that p53-dependent mechanisms block FA cells from reprogramming but also uncover a previously unrecognized defect in FA iPSC proliferation independent of p53.
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cancer and Blood Diseases Institute, Cincinnati, Ohio, USA.
authors.forename: Timothy M
authors.initials: TM
authors.lastname: Chlon
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cancer and Blood Diseases Institute, Cincinnati, Ohio, USA.
authors.forename: Elizabeth E
authors.initials: EE
authors.lastname: Hoskins
authors.affiliation: Cincinnati Children's Hospital Medical Center, Division of Developmental Biology, Cincinnati, Ohio, USA.
authors.forename: Christopher N
authors.initials: CN
authors.lastname: Mayhew
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cancer and Blood Diseases Institute, Cincinnati, Ohio, USA.
authors.forename: Kathryn A
authors.initials: KA
authors.lastname: Wikenheiser-Brokamp
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cancer and Blood Diseases Institute, Cincinnati, Ohio, USA.
authors.forename: Stella M
authors.initials: SM
authors.lastname: Davies
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cancer and Blood Diseases Institute, Cincinnati, Ohio, USA.
authors.forename: Parinda
authors.initials: P
authors.lastname: Mehta
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cancer and Blood Diseases Institute, Cincinnati, Ohio, USA.
authors.forename: Kasiani C
authors.initials: KC
authors.lastname: Myers
authors.affiliation: Cincinnati Children's Hospital Medical Center, Division of Developmental Biology, Cincinnati, Ohio, USA.
authors.forename: James M
authors.initials: JM
authors.lastname: Wells
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cancer and Blood Diseases Institute, Cincinnati, Ohio, USA Susanne.Wells@cchmc.org.
authors.forename: Susanne I
authors.initials: SI
authors.lastname: Wells
mesh_terms: D000469:Alkaline Phosphatase / Q000235:genetics / Q000378:metabolism; D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation; D049109:Cell Proliferation; D065150:Cellular Reprogramming / Q000235:genetics*; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D005199:Fanconi Anemia / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D005786:Gene Expression Regulation; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000378:metabolism / Q000473:pathology / Q000821:virology*; D015603:Keratinocytes / Q000378:metabolism / Q000473:pathology / Q000821:virology*; D000071317:Nanog Homeobox Protein; D009856:Oncogene Proteins, Viral / Q000235:genetics* / Q000378:metabolism; D061251:Primary Cell Culture; D011509:Proteoglycans / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D014161:Transduction, Genetic; D016159:Tumor Suppressor Protein p53 / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism
chemical_list: D000954:Antigens, Surface; D015415:Biomarkers; C068651:E6 protein, Human papillomavirus type 16; D018398:Homeodomain Proteins; C486780:NANOG protein, human; D000071317:Nanog Homeobox Protein; D009856:Oncogene Proteins, Viral; D011509:Proteoglycans; D034741:RNA, Small Interfering; D012097:Repressor Proteins; C107064:TRA-1-60 antigen, human; D016159:Tumor Suppressor Protein p53; D000469:Alkaline Phosphatase