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Journal of Virology, November 2008, p. 10841-10853, Vol. 82, No. 21
0022-538X/08/$08.00+0     doi:10.1128/JVI.01481-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

The E8{wedge}E2 Gene Product of Human Papillomavirus Type 16 Represses Early Transcription and Replication but Is Dispensable for Viral Plasmid Persistence in Keratinocytes{triangledown}

Michael J. Lace,1,2 James R. Anson,1 Gregory S. Thomas,2 Lubomir P. Turek,1,2 and Thomas H. Haugen1,2*

Department of Pathology, Veterans Affairs Medical Center,1 The University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa 522422

Received 15 July 2008/ Accepted 15 August 2008

A conserved E8{wedge}E2 spliced mRNA is detected in keratinocytes transfected with human papillomavirus type 16 (HPV-16) plasmid DNA. Expression of HPV-16 E8{wedge}E2 (16-E8{wedge}E2) is independent of the major early promoter, P97, and is modulated by both specific splicing events and conserved cis elements in the upstream regulatory region in a manner that differs from transcriptional regulation of other early viral genes. Mutations that disrupt the predicted 16-E8{wedge}E2 message also increase initial HPV-16 plasmid amplification 8- to 15-fold and major early gene (P97) transcription 4- to 5-fold over those of the wild type (wt). Expressing the 16-E8{wedge}E2 gene product from the cytomegalovirus (CMV) promoter represses HPV-16 early gene transcription from P97 in a dose-dependent manner, as detected by RNase protection assays. When expressed from the CMV promoter, 16-E8{wedge}E2 also inhibits the amplification of an HPV-16 plasmid and a heterologous simian virus 40 (SV40) ori plasmid that contains E2 binding sites in cis. In contrast, cotransfections with HPV-16 wt genomes that express physiologic levels of 16-E8{wedge}E2 are sufficient to repress HPV-16 plasmid amplification but are limiting and insufficient for the repression of SV40 amplification. 16-E8{wedge}E2-dependent repression of HPV-16 E1 expression is sufficient to account for this observed inhibition of initial HPV-16 plasmid amplification. Unlike with other papillomaviruses, primary human keratinocytes immortalized by the HPV-16 E8 mutant genome contain more than eightfold-higher levels of unintegrated plasmid than the wt, demonstrating that 16-E8{wedge}E2 limits the viral copy number but is not required for plasmid persistence and maintenance.

* Corresponding author. Mailing address: Department of Pathology, The University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, IA 52242. Phone: (319) 338-0581, ext. 5516. Fax: (319) 339-7178. E-mail: thomas-haugen{at}uiowa.edu

{triangledown} Published ahead of print on 27 August 2008.

Journal of Virology, November 2008, p. 10841-10853, Vol. 82, No. 21
0022-538X/08/$08.00+0     doi:10.1128/JVI.01481-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Lace, M. J., Anson, J. R., Klingelhutz, A. J., Lee, J. H., Bossler, A. D., Haugen, T. H., Turek, L. P. (2009). Human Papillomavirus (HPV) Type 18 Induces Extended Growth in Primary Human Cervical, Tonsillar, or Foreskin Keratinocytes More Effectively than Other High-Risk Mucosal HPVs. J. Virol. 83: 11784-11794 [Abstract] [Full Text]  
  • Lace, M. J., Yamakawa, Y., Ushikai, M., Anson, J. R., Haugen, T. H., Turek, L. P. (2009). Cellular factor YY1 downregulates the human papillomavirus 16 E6/E7 promoter, P97, in vivo and in vitro from a negative element overlapping the transcription-initiation site. J. Gen. Virol. 90: 2402-2412 [Abstract] [Full Text]  
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  • Lace, M. J., Anson, J. R., Klingelhutz, A. J., Harada, H., Taniguchi, T., Bossler, A. D., Haugen, T. H., Turek, L. P. (2009). Interferon-beta treatment increases human papillomavirus early gene transcription and viral plasmid genome replication by activating interferon regulatory factor (IRF)-1. Carcinogenesis 30: 1336-1344 [Abstract] [Full Text]  


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