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Epigenetic Silencing of Interferon- in Human Papillomavirus Type 16–Positive Cells

¹ Angewandte Tumorvirologie, Abteilung Virale Transformationsmechanismen, Deutsches Krebsforschungszentrum, Universität Heidelberg; ² Molekularbiologisches Labor, Universitäts-HNO-Klinik Heidelberg; ³ Molecular Alcohol Research in Gastroenterology, Universität Heidelberg; and ⁴ Angewandte Tumorvirologie, Abteilung Genomveränderungen und Karzinogenese, Deutsches Krebsforschungszentrum, Heidelberg, Germany

Requests for reprints: Frank Rösl, Angewandte Tumorvirologie, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 242, 69120 Heidelberg, Germany. Phone: 49-6221-42-4900; Fax: 49-6221-42-4902; E-mail: f.roesl{at}DKFZ.de.

We have investigated interferon- (IFN-) regulation in the context of human papillomavirus (HPV)–induced carcinogenesis using primary human foreskin keratinocytes (HFK), immortalized HFKs encoding individual oncoproteins of HPV16 (E6, E7, and E6/E7), and cervical carcinoma cells. Here, IFN- was suppressed in the presence of E6, whereas its expression was not affected in HFKs or E7-immortalized HFKs. Transcription could be reactivated after DNA demethylation but was decreased again upon drug removal. Partial reactivation could also be accomplished when E6 was knocked down, suggesting a contribution of E6 in IFN- de novo methylation. We identified a single CpG island near the transcriptional start site as being involved in selective IFN- expression. To prove the functional relevance of IFN- in building up an antiviral response, IFN- was ectopically expressed in cervical carcinoma cells where protection against vesicular stomatitis virus–mediated cytolysis could be achieved. Reconstitution of IFN- was accompanied by an increase of p53, MxA, and IFN-regulatory factors, which was reversed by knocking down either IFN- or p53 by small interfering RNA. This suggests the existence of a positive feedback loop between IFN-, p53, and components of IFN signaling pathway to maintain an antiviral state. Our in vitro findings were further corroborated in biopsy samples of cervical cancer patients, in which IFN- was also downregulated when compared with normal donor tissue. This is the first report showing an epigenetic silencing of type I IFN after HPV16 oncogene expression and revealing a novel strategy on how high-risk HPVs can abolish the innate immune response in their genuine host cells. [Cancer Res 2009;69(22):8718–25]

Key Words: innate immunity • de novo DNA methylation • antiviral activity • cervical cancer • immune escape