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Related Experiment Videos

Expression of the human papillomavirus type 11 E5A protein from the E1E4,E5 transcript

D R Brown1, T L McClowry, R A Sidner

  • 1Department of Medicine, Indiana University School of Medicine, Indianapolis, Ind., USA.

Intervirology
|August 15, 1998
PubMed
Summary

The human papillomavirus type 11 (HPV 11) E1E4,E5 transcript can express both E1E4 and E5a proteins in cells. Upstream sequences in the transcript influence HPV 11 E5a protein expression levels.

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Area of Science:

  • Virology
  • Molecular Biology

Background:

  • The human papillomavirus type 11 (HPV 11) E1E4,E5 transcript is abundant but its protein products, E5a and E5b, have not been confirmed.
  • HPV 16 E5 protein expression differs, being translated from an unspliced E2,E5 transcript, not the spliced E1E4,E5 transcript.

Purpose of the Study:

  • To determine if the HPV 11 E5a protein is expressed from the E1E4,E5 transcript.
  • To investigate the influence of upstream sequences on HPV 11 E5a protein expression.

Main Methods:

  • Green fluorescent protein (GFP) was fused to the E5a gene in various plasmid constructs.
  • COS-7 cells were transfected with constructs, and E5a-GFP expression was quantified using flow cytometry.
  • Immunofluorescence was used to detect E1E4 protein expression.

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Main Results:

  • E5a-GFP was expressed from most constructs, with varying quantities.
  • The highest E5a-GFP expression occurred when the 5' noncoding sequence (nts 714-831) was absent.
  • HPV 11 E5a-GFP was not expressed from an E2,E5a construct, unlike HPV 16.
  • Both E1E4 and E5a proteins were detected in cells transfected with a construct expressing both.

Conclusions:

  • The abundant HPV 11 E1E4,E5 transcript is a functional mRNA.
  • This transcript supports the expression of both E1E4 and E5a proteins in eukaryotic cells.
  • Sequences between nt 714 and the E5a open reading frame significantly affect E5a protein expression.