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The human papillomavirus type 11 E1E4 protein is phosphorylated in genital epithelium

J T Bryan1, A Han, K H Fife

  • 1Department of Medicine, Roudebush Veterans Administration Hospital, Indianapolis, Indiana, 46202, USA.

Virology
|March 8, 2000
PubMed

Insights

Human papillomavirus (HPV) 11 E1(wedge)E4 protein is phosphorylated by MAPK and PKA in infected cells. This phosphorylation affects viral protein localization, suggesting a role in HPV infection.

Area of Science:

  • Virology
  • Molecular Biology
  • Biochemistry

Background:

  • The E1(wedge)E4 protein is encoded by the most abundant viral transcript in human papillomavirus (HPV) 11 infections.
  • The precise function of the HPV 11 E1(wedge)E4 protein remains undetermined.
  • Potential phosphorylation sites within the E1(wedge)E4 protein suggest regulation by various protein kinases.

Purpose of the Study:

  • To investigate the phosphorylation status of the HPV 11 E1(wedge)E4 protein.
  • To identify the specific protein kinases responsible for E1(wedge)E4 phosphorylation in vitro and in vivo.
  • To determine the functional consequences of E1(wedge)E4 phosphorylation on its intracellular localization.

Main Methods:

  • Production of a maltose binding protein (MBP) fusion of HPV 11 E1(wedge)E4 in E. coli for in vitro kinase assays.
  • Phosphoamino acid analysis and liquid chromatography-mass spectrometry to identify phosphorylated residues.
  • Site-directed mutagenesis to create E1(wedge)E4 phosphoacceptor mutants and analysis of their intracellular distribution in HPV 11-infected xenografts.

Main Results:

  • Mitogen-activated protein kinase (MAPK) and cAMP-dependent protein kinase (PKA) were identified as the kinases phosphorylating E1(wedge)E4 in vitro.
  • MAPK phosphorylates E1(wedge)E4 at threonine 53, while PKA phosphorylates at threonine 36 and serine 44.
  • In vivo analysis of HPV 11-infected xenografts confirmed serine and threonine phosphorylation of E1(wedge)E4, consistent with MAPK and PKA activity. The MAPK mutant exhibited altered intracellular localization compared to wild-type.

Conclusions:

  • The HPV 11 E1(wedge)E4 protein undergoes phosphorylation both in vitro and in vivo.
  • MAPK and PKA are key kinases involved in the phosphorylation of E1(wedge)E4 in HPV-infected cells.
  • Phosphorylation, particularly by MAPK, influences the intracellular distribution of E1(wedge)E4, suggesting a role in viral pathogenesis.

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