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PMID: 16987971 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't

Binding and nuclear relocalization of protein kinase R by human cytomegalovirus TRS1.

Journal of virology ·Vol. 80 ·No. 23 ·2006-12-00 ·Pages 11817-26

Hakki M, Marshall EE, De Niro KL, Geballe AP

Abstract

The human cytomegalovirus (HCMV) TRS1 and IRS1 genes block the phosphorylation of the alpha subunit of eukaryotic initiation factor 2 (eIF2alpha) and the consequent shutoff of cellular protein synthesis that occur during infection with vaccinia virus (VV) deleted of the double-stranded RNA binding protein gene E3L (VVDeltaE3L). To further define the underlying mechanism, we first evaluated the effect of pTRS1 on protein kinase R (PKR), the double-stranded RNA (dsRNA)-dependent eIF2alpha kinase. Immunoblot analyses revealed that pTRS1 expression in the context of a VVDeltaE3L recombinant decreased levels of PKR in the cytoplasm and increased its levels in the nucleus of infected cells, an effect not seen with wild-type VV or a VVDeltaE3L recombinant virus expressing E3L. This effect of pTRS1 was confirmed by visualizing the nuclear relocalization of PKR-EGFP expressed by transient transfection. PKR present in both the nuclear and cytoplasmic fractions was nonphosphorylated, indicating that it was unactivated when TRS1 was present. PKR also accumulated in the nucleus during HCMV infection as determined by indirect immunofluorescence and immunoblot analysis. Binding assays revealed that pTRS1 interacted with PKR in mammalian cells and in vitro. This interaction required the same carboxy-terminal region of pTRS1 that is necessary to rescue VVDeltaE3L replication in HeLa cells. The carboxy terminus of pIRS1 was also required for rescue of VVDeltaE3L and for mediating an interaction of pIRS1 with PKR. These results suggest that these HCMV genes directly interact with PKR and inhibit its activation by sequestering it in the nucleus, away from both its activator, cytoplasmic dsRNA, and its substrate, eIF2alpha.

MeSH Terms
Cytomegalovirus/genetics,metabolism,physiology HeLa Cells Humans Protein Kinases/genetics,metabolism RNA, Double-Stranded/metabolism RNA-Binding Proteins/chemistry,genetics,metabolism Viral Proteins/metabolism
Chemicals
RNA, Double-Stranded RNA-Binding Proteins Viral Proteins Protein Kinases
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Hakki Morgan
Division of Human Biology, Fred Hutchinson Cancer Research Center, 1100 Fairview Ave. N., Seattle, WA 98109, USA.
Marshall Emily E
De Niro Katherine L
Geballe Adam P
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Article Info
Journal
Journal of virology
Abbr.
J Virol
ISSN
0022-538X
DOI
10.1128/JVI.00957-06
Published
2006-12-00
Epub
2006-00-20
Pages
11817-26
Language
English
Region
United States
NLM ID
0113724
PMCID
PMC1642616
Subset
IM
Grants
NIAID NIH HHS · K08 AI058089 · United States
NIAID NIH HHS · R01 AI 026672 · United States
NCI NIH HHS · T32 CA 09229 · United States
NCI NIH HHS · T32 CA009229 · United States
NIAID NIH HHS · R01 AI026672 · United States
NIAID NIH HHS · K08 AI 058089 · United States
Corrections
ErratumIn
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External Links
PubMed source DOI: 10.1128/JVI.00957-06 PMC Full Text View journal details
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