title: Identification of human papillomavirus (HPV) 16 DNA integration and the ensuing patterns of methylation in HPV-associated head and neck squamous cell carcinoma cell lines.
abstract: Recent studies showed that human papillomavirus (HPV) integration contributes to the genomic instability seen in HPV-associated head and neck squamous cell carcinoma (HPV-HNSCC). However, the epigenetic alterations induced after HPV integration remains unclear. To identify the molecular details of HPV16 DNA integration and the ensuing patterns of methylation in HNSCC, we performed next-generation sequencing using a target-enrichment method for the effective identification of HPV16 integration breakpoints as well as the characterization of genomic sequences adjacent to HPV16 integration breakpoints with three HPV16-related HNSCC cell lines. The DNA methylation levels of the integrated HPV16 genome and that of the adjacent human genome were also analyzed by bisulfite pyrosequencing. We found various integration loci, including novel integration sites. Integration loci were located predominantly in the intergenic region, with a significant enrichment of the microhomologous sequences between the human and HPV16 genomes at the integration breakpoints. Furthermore, various levels of methylation within both the human genome and the integrated HPV genome at the integration breakpoints in each integrant were observed. Allele-specific methylation analysis suggested that the HPV16 integrants remained hypomethylated when the flanking host genome was hypomethylated. After integration into highly methylated human genome regions, however, the HPV16 DNA became methylated. In conclusion, we found novel integration sites and methylation patterns in HPV-HNSCC using our unique method. These findings may provide insights into understanding of viral integration mechanism and virus-associated carcinogenesis of HPV-HNSCC.
authors.affiliation: Department of Biology and Function in Head and Neck, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Hatano
authors.affiliation: Department of Biology and Function in Head and Neck, Yokohama City University Graduate School of Medicine, Yokohama, Japan.|Department of Otorhinolaryngology - Head and Neck Surgery, Yokohama City University, School of Medicine, Yokohama, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Sano
authors.affiliation: Department of Biology and Function in Head and Neck, Yokohama City University Graduate School of Medicine, Yokohama, Japan.|Department of Head and Neck Surgery, The University of Texas MD Anderson Cancer Center, Houston, TX.
authors.forename: Hideaki
authors.initials: H
authors.lastname: Takahashi
authors.affiliation: Department of Biology and Function in Head and Neck, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Hyakusoku
authors.affiliation: Department of Biology and Function in Head and Neck, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Isono
authors.affiliation: Department of Biology and Function in Head and Neck, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Shoko
authors.initials: S
authors.lastname: Shimada
authors.affiliation: Department of Biology and Function in Head and Neck, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Kae
authors.initials: K
authors.lastname: Sawakuma
authors.affiliation: Department of Biology and Function in Head and Neck, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Takada
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, St. Marianna University School of Medicine, Kawasaki, Japan.
authors.forename: Ritsuko
authors.initials: R
authors.lastname: Oikawa
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, St. Marianna University School of Medicine, Kawasaki, Japan.
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Watanabe
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, St. Marianna University School of Medicine, Kawasaki, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Yamamoto
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, St. Marianna University School of Medicine, Kawasaki, Japan.
authors.forename: Fumio
authors.initials: F
authors.lastname: Itoh
authors.affiliation: Department of Head and Neck Surgery, The University of Texas MD Anderson Cancer Center, Houston, TX.
authors.forename: Jeffrey N
authors.initials: JN
authors.lastname: Myers
authors.affiliation: Department of Biology and Function in Head and Neck, Yokohama City University Graduate School of Medicine, Yokohama, Japan.|Department of Otorhinolaryngology - Head and Neck Surgery, Yokohama City University, School of Medicine, Yokohama, Japan.
authors.forename: Nobuhiko
authors.initials: N
authors.lastname: Oridate
mesh_terms: D000483:Alleles; D002294:Carcinoma, Squamous Cell / Q000235:genetics / Q000821:virology*; D045744:Cell Line, Tumor; D019175:DNA Methylation; D004279:DNA, Viral / Q000235:genetics*; D018450:Disease Progression; D016679:Genome, Viral; D006258:Head and Neck Neoplasms / Q000235:genetics / Q000821:virology*; D052162:Human papillomavirus 16 / Q000502:physiology*; D006801:Humans; D020084:Long Interspersed Nucleotide Elements; D008297:Male; D008875:Middle Aged; D009856:Oncogene Proteins, Viral / Q000235:genetics; D030361:Papillomavirus Infections / Q000235:genetics / Q000821:virology; D000077195:Squamous Cell Carcinoma of Head and Neck; D016662:Virus Integration
chemical_list: D004279:DNA, Viral; D009856:Oncogene Proteins, Viral
keywords: DNA integration; epigenome; head and neck squamous cell carcinoma (HNSCC); human papillomavirus (HPV)