Skip to main content

Analysis of the Regulation of Viral Transcription

  • Protocol
Human Papillomaviruses

Part of the book series: Methods in Molecular Medicine ((MIMM,volume 119))

  • 1262 Accesses

Summary

Despite the small genomes and number of genes of papillomaviruses, regulation of their transcription is very complex and governed by numerous transcription factors, cis-responsive elements, and epigenetic phenomena. This chapter describes the strategies of how one can approach a systematic analysis of these factors, elements, and mechanisms. From the numerous different techniques useful for studying transcription, we describe in detail three selected protocols of approaches that have been relevant in shaping our knowledge of human papillomavirus transcription. These are DNAse I protection (“footprinting”) for location of transcription-factor binding sites, electrophoretic mobility shifts (“gelshifts”) for analysis of bound transcription factors, and bisulfite sequencing for analysis of DNA methylation as a prerequisite for epigenetic transcriptional regulation.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Bernard, H. U. (2002) Gene expression of genital human papillomaviruses and potential antiviral approaches. Antiviral Therapy 7, 219–237.

    PubMed  CAS  Google Scholar 

  2. Hummel, M., Hudson, J. B., and Laimins, L. A. (1992) Differentiation-induced and constitutive transcription of human papillomavirus type 31b in cell lines containing viral episomes. J. Virol. 66, 6070–6080.

    PubMed  CAS  Google Scholar 

  3. Meyers, C., Mayer, T. J., and Ozbun, M. A. (1997) Synthesis of infectious human papillomavirus type 18 in differentiating epithelium transfected with viral DNA. J. Virol. 71, 7381–7386.

    PubMed  CAS  Google Scholar 

  4. Stoler, M. and Broker, T. H. (1986) In situ hybridization detection of human papillomavirus DNAs and messenger RNAs in genital condylomas and cervical carcinoma. Human Pathol. 17, 1250–1258.

    Article  CAS  Google Scholar 

  5. Sailaja, G., Watts, R. M., and Bernard, H. U. (1999) Many different papillomaviruses have low transcriptional activity in spite of strong epithelial specific enhancers. J. Gen. Virol. 80, 1715–1724.

    PubMed  CAS  Google Scholar 

  6. Chan, W. K., Chong, T., Bernard, H. U., and Klock, G. (1990) Two AP1 sites in the long control region of human papillomavirus type 16 lead to phorbolester stimulation of the viral E6/E7 promoter. Nucleic Acids Res. 18, 763–769.

    Article  PubMed  CAS  Google Scholar 

  7. Ozbun, M. A. and Meyers, C. (1997) Characterization of late gene transcripts expressed during vegetative replication of human papillomavirus type 31b. J. Virol. 71, 5161–5172.

    PubMed  CAS  Google Scholar 

  8. Tan, S. H., Leong, L. E. C., Walker, P. A., and Bernard, H. U. (1994) The human papillomavirus type 16 transcription factor E2 binds with low cooperativity to two flanking binding sites and represses the E6 promoter through displacement of Sp1 and TFIID. J. Virol. 68, 6411–6420.

    PubMed  CAS  Google Scholar 

  9. Ai, W., Narahari, J., and Roman, A. (2000) Ying yang 1 negatively regulates the differentiation-specific E1 promoter of human papillomavirus 6. J. Virol. 74, 5198–5205.

    Article  PubMed  CAS  Google Scholar 

  10. Ai, W., Toussaint, E., and Roman, A. (1999) CCAAT displacement protein binds to and negatively regulates human papillomavirus type 6 E6, E7, and E1 promoters. J. Virol. 73, 4220–4229.

    PubMed  CAS  Google Scholar 

  11. O’Connor, M. J., Stünkel, W., Koh, C. H., Zimmermann, H., and Bernard, H. U. (2000) The differentiation-specific factor CDP/Cut represses transcription and replication of human papillomaviruses. J. Virol. 74, 401–410.

    Article  PubMed  Google Scholar 

  12. Stünkel, W., Huang, Z., Tan, S. H., O’Connor, M, and Bernard, H. U. (2000) Nuclear matrix attachment regions of human papillomavirus-16 repress or activate the E6 promoter depending on the physical state of the viral DNA. J. Virol. 74, 2489–2501.

    Article  PubMed  Google Scholar 

  13. Chong, T., Apt, D., Gloss, B., Isa, M., and Bernard, H. U. (1991) The enhancer of human papillomavirus-16: Binding sites for the ubiquitous transcription factors oct-1, NFA, TEF-2, NFI and AP1 participate in the epithelial specific transcription. J.Virol. 65, 5933–5943.

    PubMed  CAS  Google Scholar 

  14. Gloss, B., Chong, T., and Bernard, H. U. (1989) Numerous nuclear proteins bind the long control region of human papillomavirus type 16: A subset of 6 of 23 DNAseI-protected segments coincides with the location of the cell-type-specific enhancer. J.Virol. 63, 1142–1152.

    PubMed  CAS  Google Scholar 

  15. Stünkel, W. and Bernard, H. U. (1999) The chromatin structure of the long control region of human papillomavirus type 16 represses viral oncoprotein expression. J. Virol. 73, 1918–1930.

    PubMed  Google Scholar 

  16. Badal, V., Chuang, L. S. H., Badal, S., et al. (2003) CpG methylation of human papillomavirus-16 DNA in cervical cancer cell lines and in clinical specimens: genomic hypomethylation correlates with carcinogenic progression. J. Virol. 77, 6227–6234.

    Article  PubMed  Google Scholar 

  17. Dignam, J. D., Lebovitz, R. M., and Roeder, R. G. (1983) Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 11, 1475–1489.

    Article  PubMed  CAS  Google Scholar 

  18. Wildeman, A. G., Sassone-Corsi, P., Grundstrom, T., Zenke, M., and Chambon, P. (1984) Stimulation of in vitro transcription from the SV40 early promoter by the enhancer involves a specific trans-acting factor. EMBO J. 3, 3129–3133.

    PubMed  CAS  Google Scholar 

  19. Frommer, M., McDonald, L. E., Millar, D. S., et al. (1992) A genomic sequencing protocol that yields a positive display of 5-methylcytosine residues in individual DNA strands. Proc. Natl. Acad. Sci. USA 89, 1827–1831.

    Article  PubMed  CAS  Google Scholar 

  20. Millar, D. S., Warnecke, P. M., Melki, J. R., and Clark, S. J. (2002) Methylation sequencing from limiting DNA: embryonic, fixed, and microdissected cells. Methods 27, 108–113.

    Article  PubMed  CAS  Google Scholar 

  21. Kim, K., Garner-Hamrick, P. A., Fisher, C., Lee, D., and Lambert, P. F. (2003) Methylation patterns of papillomavirus DNA, its influence on E2 function, and implications in viral infection. J. Virol. 77, 12,450–12,459.

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2005 Humana Press Inc.

About this protocol

Cite this protocol

Gloss, B., Kalantari, M., Bernard, HU. (2005). Analysis of the Regulation of Viral Transcription. In: Davy, C., Doorbar, J. (eds) Human Papillomaviruses. Methods in Molecular Medicine, vol 119. Humana Press. https://doi.org/10.1385/1-59259-982-6:261

Download citation

  • DOI: https://doi.org/10.1385/1-59259-982-6:261

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-373-2

  • Online ISBN: 978-1-59259-982-0

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics