N-Terminal Phosphorylation Sites of Herpes Simplex Virus 1 ICP0 Differentially Regulate Its Activities and Enhance Viral Replication

Heba Mostafa, Thornton W. Thompson, David J. Davido

Research output: Contribution to journalArticle

Abstract

The herpes simplex virus 1 (HSV-1) infected cell protein 0 (ICP0) is an immediate-early phosphoprotein that transactivates viral gene expression. Evidence suggests that phosphorylation regulates the functions of ICP0, and three regions (termed regions I, II, and III) in the protein are known to be phosphorylated. Mutation of the putative phosphorylation sites within region I, termed Phos 1, which lies in the N-terminal portion of ICP0, impairs the E3 ubiquitin (Ub) ligase and ND10-disrupting activities of ICP0 in cell culture and diminishes viral replication. To identify the specific phosphorylation site(s) or residues responsible for the phenotypes observed with Phos 1, individual residues within region I were mutated to alanine (S224A, T226A, T231A, and T232A) and one double mutant S224A/T226A was constructed. Tissue culture studies demonstrated that the S224A, S224A/ T226A, T231A, and T232A mutants were unable to dissociate the cellular protein PML from ND10 and that the S224/T226A mutant was defective in its ability to dissociate the cellular protein Sp100 from ND10. Additionally, the transactivation activity of ICP0 was impaired in the S224A and S224A/T226A mutants. The S224A and S224A/T226A mutant forms were more stable than wild-type ICP0, suggesting that their ability to autoubiquitinate was limited. Moreover, one ICP0 ubiquitination target, USP-7, was also more stable after infection with these two mutants. Lastly, the replication of the S224A and S224A/T226A mutant viruses was reduced in cell culture and in vivo. Overall, our data suggest that specific phosphorylation sites within region I differentially regulate the activities of ICP0, which are required for efficient viral replication.

Original languageEnglish (US)
Pages (from-to)2109-2119
Number of pages11
JournalJournal of virology
Volume87
Issue number4
DOIs
StatePublished - Feb 1 2013
Externally publishedYes

Fingerprint

Human herpesvirus 1
virus replication
phosphorylation
Phosphorylation
Proteins
mutants
proteins
cells
Cell Culture Techniques
cell culture
Human herpesvirus 1 Vmw110 protein
Ubiquitin-Protein Ligases
Viral Genes
Ubiquitination
Phosphoproteins
phosphoproteins
ubiquitin-protein ligase
transcriptional activation
Alanine
Transcriptional Activation

ASJC Scopus subject areas

  • Immunology
  • Virology

Cite this

N-Terminal Phosphorylation Sites of Herpes Simplex Virus 1 ICP0 Differentially Regulate Its Activities and Enhance Viral Replication. / Mostafa, Heba; Thompson, Thornton W.; Davido, David J.

In: Journal of virology, Vol. 87, No. 4, 01.02.2013, p. 2109-2119.

Research output: Contribution to journalArticle

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abstract = "The herpes simplex virus 1 (HSV-1) infected cell protein 0 (ICP0) is an immediate-early phosphoprotein that transactivates viral gene expression. Evidence suggests that phosphorylation regulates the functions of ICP0, and three regions (termed regions I, II, and III) in the protein are known to be phosphorylated. Mutation of the putative phosphorylation sites within region I, termed Phos 1, which lies in the N-terminal portion of ICP0, impairs the E3 ubiquitin (Ub) ligase and ND10-disrupting activities of ICP0 in cell culture and diminishes viral replication. To identify the specific phosphorylation site(s) or residues responsible for the phenotypes observed with Phos 1, individual residues within region I were mutated to alanine (S224A, T226A, T231A, and T232A) and one double mutant S224A/T226A was constructed. Tissue culture studies demonstrated that the S224A, S224A/ T226A, T231A, and T232A mutants were unable to dissociate the cellular protein PML from ND10 and that the S224/T226A mutant was defective in its ability to dissociate the cellular protein Sp100 from ND10. Additionally, the transactivation activity of ICP0 was impaired in the S224A and S224A/T226A mutants. The S224A and S224A/T226A mutant forms were more stable than wild-type ICP0, suggesting that their ability to autoubiquitinate was limited. Moreover, one ICP0 ubiquitination target, USP-7, was also more stable after infection with these two mutants. Lastly, the replication of the S224A and S224A/T226A mutant viruses was reduced in cell culture and in vivo. Overall, our data suggest that specific phosphorylation sites within region I differentially regulate the activities of ICP0, which are required for efficient viral replication.",
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