title: Expression of betapapillomavirus oncogenes increases the number of keratinocytes with stem cell-like properties.
abstract: Human papillomaviruses (HPV) of genus Betapapillomavirus (betaPV) are associated with nonmelanoma skin cancer development in epidermodysplasia verruciformis (EV) and immunosuppressed patients. Epidemiological and molecular studies suggest a carcinogenic activity of betaPV during early stages of cancer development. Since viral oncoproteins delay and perturb keratinocyte differentiation, they may have the capacity to either retain or confer a "stem cell-like" state on oncogene-expressing cells. The aim of this study was to determine (i) whether betaPV alters the expression of cell surface markers, such as CD44 and epithelial cell adhesion molecule (EpCAM), that have been associated with epithelial stemness, and (ii) whether this confers functional stem cell-like properties to human cutaneous keratinocytes. Fluorescence-activated cell sorter (FACS) analysis revealed an increase in the number of cells with high CD44 and EpCAM expression in keratinocyte cultures expressing HPV type 8 (HPV8) oncogenes E2, E6, and E7. Particularly through E7 expression, a distinct increase in clonogenicity and in the formation and size of tumor spheres was observed, accompanied by reduction of the epithelial differentiation marker Calgranulin B. These stem cell-like properties could be attributed to the pool of CD44(high) EpCAM(high) cells, which was increased within the E7 cultures of HPV5, -8, and -20. Enhanced EpCAM levels were present in organotypic skin cultures of primary keratinocytes expressing E7 of the oncogenic HPV types HPV5, -8, and -16 and in clinical samples from EV patients. In conclusion, our data show that betaPV may increase the number of stem cell-like cells present during early carcinogenesis and thus enable the persistence and accumulation of DNA damage necessary to generate malignant stem cells.
authors.affiliation: Institute of Virology, University of Cologne, Cologne, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Hufbauer
authors.forename: Adrian
authors.initials: A
authors.lastname: Biddle
authors.forename: Cinzia
authors.initials: C
authors.lastname: Borgogna
authors.forename: Marisa
authors.initials: M
authors.lastname: Gariglio
authors.forename: John
authors.initials: J
authors.lastname: Doorbar
authors.forename: Alan
authors.initials: A
authors.lastname: Storey
authors.forename: Herbert
authors.initials: H
authors.lastname: Pfister
authors.forename: Ian
authors.initials: I
authors.lastname: Mackenzie
authors.forename: Baki
authors.initials: B
authors.lastname: Akgül
mesh_terms: D000951:Antigens, Neoplasm / Q000235:genetics / Q000378:metabolism; D017209:Apoptosis; D052686:Betapapillomavirus / Q000472:pathogenicity; D015153:Blotting, Western; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D000071858:Epithelial Cell Adhesion Molecule; D005455:Fluorescent Antibody Technique; D006801:Humans; D018960:Hyaluronan Receptors / Q000235:genetics / Q000378:metabolism; D015603:Keratinocytes / Q000378:metabolism / Q000473:pathology / Q000821:virology*; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000378:metabolism*; D030361:Papillomavirus Infections / Q000378:metabolism / Q000473:pathology / Q000821:virology*; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012871:Skin Diseases / Q000378:metabolism / Q000473:pathology / Q000821:virology*; D013234:Stem Cells / Q000378:metabolism / Q000473:pathology / Q000821:virology*
chemical_list: D000951:Antigens, Neoplasm; C497483:CD44 protein, human; D015815:Cell Adhesion Molecules; C499379:EPCAM protein, human; D000071858:Epithelial Cell Adhesion Molecule; D018960:Hyaluronan Receptors; D009856:Oncogene Proteins, Viral; D012333:RNA, Messenger; C066076:oncogene protein E7, Human papillomavirus type 8