title: Differentiation-Dependent KLF4 Expression Promotes Lytic Epstein-Barr Virus Infection in Epithelial Cells.
abstract: Epstein-Barr virus (EBV) is a human herpesvirus associated with B-cell and epithelial cell malignancies. EBV lytically infects normal differentiated oral epithelial cells, where it causes a tongue lesion known as oral hairy leukoplakia (OHL) in immunosuppressed patients. However, the cellular mechanism(s) that enable EBV to establish exclusively lytic infection in normal differentiated oral epithelial cells are not currently understood. Here we show that a cellular transcription factor known to promote epithelial cell differentiation, KLF4, induces differentiation-dependent lytic EBV infection by binding to and activating the two EBV immediate-early gene (BZLF1 and BRLF1) promoters. We demonstrate that latently EBV-infected, telomerase-immortalized normal oral keratinocyte (NOKs) cells undergo lytic viral reactivation confined to the more differentiated cell layers in organotypic raft culture. Furthermore, we show that endogenous KLF4 expression is required for efficient lytic viral reactivation in response to phorbol ester and sodium butyrate treatment in several different EBV-infected epithelial cell lines, and that the combination of KLF4 and another differentiation-dependent cellular transcription factor, BLIMP1, is highly synergistic for inducing lytic EBV infection. We confirm that both KLF4 and BLIMP1 are expressed in differentiated, but not undifferentiated, epithelial cells in normal tongue tissue, and show that KLF4 and BLIMP1 are both expressed in a patient-derived OHL lesion. In contrast, KLF4 protein is not detectably expressed in B cells, where EBV normally enters latent infection, although KLF4 over-expression is sufficient to induce lytic EBV reactivation in Burkitt lymphoma cells. Thus, KLF4, together with BLIMP1, plays a critical role in mediating lytic EBV reactivation in epithelial cells.
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America; Cellular and Molecular Biology Graduate Program, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Dhananjay M
authors.initials: DM
authors.lastname: Nawandar
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America; Cellular and Molecular Biology Graduate Program, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Anqi
authors.initials: A
authors.lastname: Wang
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Kathleen
authors.initials: K
authors.lastname: Makielski
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Denis
authors.initials: D
authors.lastname: Lee
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Shidong
authors.initials: S
authors.lastname: Ma
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Barlow
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America; Cellular and Molecular Biology Graduate Program, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Jessica
authors.initials: J
authors.lastname: Reusch
authors.affiliation: Department of Microbiology and Immunology, Center for Molecular and Tumor Virology and Feist-Weiller Cancer Center, Louisiana State University Health Sciences Center, Shreveport, Louisiana, United States of America.
authors.forename: Ru
authors.initials: R
authors.lastname: Jiang
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America; Medical Microbiology and Immunology Graduate Program, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Coral K
authors.initials: CK
authors.lastname: Wille
authors.affiliation: Department of Orofacial Sciences, School of Dentistry, University of California, San Francisco, San Francisco, California, United States of America.
authors.forename: Deborah
authors.initials: D
authors.lastname: Greenspan
authors.affiliation: Department of Orofacial Sciences, School of Dentistry, University of California, San Francisco, San Francisco, California, United States of America.
authors.forename: John S
authors.initials: JS
authors.lastname: Greenspan
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Janet E
authors.initials: JE
authors.lastname: Mertz
authors.affiliation: Department of Microbiology and Immunology, Center for Molecular and Tumor Virology and Feist-Weiller Cancer Center, Louisiana State University Health Sciences Center, Shreveport, Louisiana, United States of America.
authors.forename: Lindsey
authors.initials: L
authors.lastname: Hutt-Fletcher
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America; Department of Medicine, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Eric C
authors.initials: EC
authors.lastname: Johannsen
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Paul F
authors.initials: PF
authors.lastname: Lambert
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America; Department of Medicine, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.
authors.forename: Shannon C
authors.initials: SC
authors.lastname: Kenney
mesh_terms: D000328:Adult; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line; D047369:Chromatin Immunoprecipitation; D004847:Epithelial Cells / Q000473:pathology / Q000821:virology*; D020031:Epstein-Barr Virus Infections / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D054884:Host-Pathogen Interactions / Q000502:physiology; D006801:Humans; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000378:metabolism*; D061346:Laser Capture Microdissection; D017733:Leukoplakia, Hairy / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D000074462:Positive Regulatory Domain I-Binding Factor 1; D012097:Repressor Proteins / Q000378:metabolism*; D014775:Virus Activation / Q000502:physiology*; D017735:Virus Latency / Q000502:physiology
chemical_list: C000715467:KLF4 protein, human; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D012097:Repressor Proteins; C087042:PRDM1 protein, human; D000074462:Positive Regulatory Domain I-Binding Factor 1