title: Depletion of HPV16 early genes induces autophagy and senescence in a cervical carcinogenesis model, regardless of viral physical state.
abstract: In cervical carcinomas, high-risk human papillomavirus (HR-HPV) may be integrated into host chromosomes or remain extra-chromosomal (episomal). We used the W12 cervical keratinocyte model to investigate the effects of HPV16 early gene depletion on in vitro cervical carcinogenesis pathways, particularly effects shared by cells with episomal versus integrated HPV16 DNA. Importantly, we were able to study the specific cellular consequences of viral gene depletion by using short interfering RNAs known not to cause phenotypic or transcriptional off-target effects in keratinocytes. We found that while cervical neoplastic progression in vitro was characterized by dynamic changes in HPV16 transcript levels, viral early gene expression was required for cell survival at all stages of carcinogenesis, regardless of viral physical state, levels of early gene expression or histology in organotypic tissue culture. Moreover, HPV16 early gene depletion induced changes in host gene expression that were common to both episome-containing and integrant-containing cells. In particular, we observed up-regulation of autophagy genes, associated with enrichment of senescence and innate immune-response pathways, including the senescence-associated secretory phenotype (SASP). In keeping with these observations, HPV16 early gene depletion induced autophagy in both episome-containing and integrant-containing W12 cells, as evidenced by the appearance of autophagosomes, punctate expression of the autophagy marker LC3, conversion of LC3B-I to LC3B-II, and reduced levels of the autophagy substrate p62. Consistent with the reported association between autophagy and senescence pathways, HPV16 early gene depletion induced expression of the senescence marker beta-galactosidase and increased secretion of the SASP-related protein IGFBP3. Together, these data indicate that depleting HR-HPV early genes would be of potential therapeutic benefit in all cervical carcinogenesis pathways, regardless of viral physical state. In addition, the senescence/SASP response associated with autophagy induction may promote beneficial immune effects in bystander cells.
authors.affiliation: Department of Pathology, University of Cambridge, CB2 1QP, UK.
authors.forename: Jennifer E
authors.initials: JE
authors.lastname: Hanning
authors.forename: Harpreet K
authors.initials: HK
authors.lastname: Saini
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Murray
authors.forename: Maria M
authors.initials: MM
authors.lastname: Caffarel
authors.forename: Stijn
authors.initials: S
authors.lastname: van Dongen
authors.forename: Dawn
authors.initials: D
authors.lastname: Ward
authors.forename: Emily M
authors.initials: EM
authors.lastname: Barker
authors.forename: Cinzia G
authors.initials: CG
authors.lastname: Scarpini
authors.forename: Ian J
authors.initials: IJ
authors.lastname: Groves
authors.forename: Margaret A
authors.initials: MA
authors.lastname: Stanley
authors.forename: Anton J
authors.initials: AJ
authors.lastname: Enright
authors.forename: Mark R
authors.initials: MR
authors.lastname: Pett
authors.forename: Nicholas
authors.initials: N
authors.lastname: Coleman
mesh_terms: D001343:Autophagy / Q000235:genetics; D045744:Cell Line, Tumor; D002472:Cell Transformation, Viral / Q000235:genetics*; D016922:Cellular Senescence / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D015967:Gene Expression Regulation, Viral; D052162:Human papillomavirus 16 / Q000235:genetics*; D006801:Humans; D009856:Oncogene Proteins, Viral / Q000235:genetics; D050725:Papillomavirus E7 Proteins / Q000235:genetics; D030361:Papillomavirus Infections / Q000150:complications / Q000821:virology*; D010641:Phenotype; D010957:Plasmids; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D012367:RNA, Viral / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics; D013997:Time Factors; D014162:Transfection; D002583:Uterine Cervical Neoplasms / Q000235:genetics / Q000473:pathology* / Q000821:virology*; D016662:Virus Integration
chemical_list: C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D012333:RNA, Messenger; D012367:RNA, Viral; D012097:Repressor Proteins; C059731:oncogene protein E7, Human papillomavirus type 16
keywords: HPV16; autophagy; cervix; senescence; squamous cell carcinoma