Protein kinase B/Akt is present in activated form throughout the entire replicative cycle of deltaU(S)3 mutant virus but only at early times after infection with wild-type herpes simplex virus 1.

Benetti L, Roizman B.

The Marjorie B. Kovler Viral Oncology Laboratories, University of Chicago, 910 East 58th Street, Chicago, IL 60637, USA.

The product of the herpes simplex virus 1 (HSV-1) US3 gene is a multifunctional serine-threonine protein kinase that can block apoptosis induced by proapoptotic cellular proteins, exogenous agents, or replication-defective viruses. Earlier studies showed that the U(S)3 kinase activates and functionally overlaps cellular protein kinase A (PKA). In this study we examined the status of phosphatidylinositol 3-kinase [PI3K] and of its effector, protein kinase B/Akt (PKB/Akt), a component of a major pathway of mammalian antiapoptotic signaling systems. We report the following. (i) Infection of target cells with HSV-1 induces transient phosphorylation of serine 473 of PKB/Akt early in infection, with a mechanism that is dependent on PI3K. Inhibition of PI3K induced apoptosis in mock-infected or deltaU(S)3 mutant-virus-infected but not in wild-type-virus-infected cells and reduced the accumulation of specific viral gene products, including the U(S)3 protein kinase, but had a marginal effect on virus yields. (ii) At later times after infection, the total amounts of PKB/Akt decreased and phosphorylated PKB/Akt forms disappeared in a U(S)3-dependent and protein phosphatase 2A-independent manner. (iii) Activation of PKA by forskolin did not mediate significant dephosphorylation of PKB/Akt. Our results are consistent with the model that PKB/Akt is activated early in infection and acts to block apoptosis in infected cells prior to the accumulation of U(S)3 protein kinase and that it persists and continues to function as an antiapoptotic protein in the absence of U(S)3 but becomes redundant or even inimical once U(S)3 protein kinase accumulates in effective amounts.

PMID: 16537601 [PubMed - in process]

71792 [PubMed - in process]

2007/8-27/ 31

J Virol. 2006 Apr;80(8):3752-64.

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U(S)3 and U(S)3.5 protein kinases of herpes simplex virus 1 differ with respect to their functions in blocking apoptosis and in virion maturation and egress.

Poon AP, Benetti L, Roizman B.

The Marjorie B. Kovler Viral Oncology Laboratories, The University of Chicago, 910 East 58th Street, Chicago, Illinois 60637, USA.

Previously, we reported that the U(S)3 protein kinase blocks apoptosis, that it activates protein kinase A (PKA), that activation of PKA blocks apoptosis in cells infected with a U(S)3 deletion mutant, and that an overlapping transcriptional unit encodes a truncated kinase designated U(S)3.5. Here, we report the properties of the kinases based on comparisons of herpes simplex virus and baculoviruses expressing U(S)3 or U(S)3.5 kinase. Specifically, we report the following. (i) Both kinases mediate the phosphorylation of HDAC1, HDAC2, and the PKA regulatory IIalpha subunit in the absence of other viral proteins. (ii) Both enzymes mediate the phosphorylation of largely identical sets of proteins carrying the phosphorylation consensus site of PKA, but only U(S)3 blocks apoptosis, suggesting that it is U(S)3 and not PKA that is responsible for the phosphorylation of the proteins bearing the shared consensus phosphorylation site and the antiapoptotic activity. (iii) Both kinases cofractionate with mitochondria. Immune depletion of the U(S)3 and U(S)3.5 kinases from the cytoplasm removed the kinases from the supernatant fraction, but not from the mitochondrial fraction, and therefore, if the antiapoptotic activity of the U(S)3 kinase is expressed in mitochondria, the localization signal and the antiapoptotic functions are located on different parts of the protein. (iv) The U(S)3 protein kinase is required for the translocation of virus particles from the nucleus. Although the U(L)31 protein is phosphorylated in cells infected with the mutant expressing U(S)3.5 kinase, the release of virus particles from nuclei was impeded in some cells, suggesting that the U(S)3 kinase affects the modification of the nuclear membrane more efficiently than the U(S)3.5 kinase.

PMID: 165

J Virol. 2006 Apr;80(7):3341-8.

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