2-Aminopurine Selectively Inhibits the Induction of β-Interferon, c-
            <i>fos</i>
            , and c-
            <i>myc</i>
            Gene Expression

Zinn, Kai; Keller, Andrew; Whittemore, Lisa-Anne; Maniatis, Tom

doi:10.1126/science.3281258

Cited by 227 publications

(120 citation statements)

References 19 publications

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“…Additionally, there is indirect evidence implicating PKR in the transcriptional up-regulation of select genes in response to viral infection or dsRNA (75), and PKR has been demonstrated to directly interact with several proteins present in the nucleus that may affect regulation of the cell cycle (22,23,55). While the functional significance of these interactions is unclear, PKR relocalization to the nucleus during HCMV infection may serve a role distinct from its role in the eIF2␣ pathway.…”

Section: Discussionmentioning

confidence: 99%

Binding and Nuclear Relocalization of Protein Kinase R by Human Cytomegalovirus TRS1

Hakki

Marshall²,

Niro³

et al. 2006

J Virol

103

100

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The human cytomegalovirus (HCMV) TRS1 and IRS1 genes block the phosphorylation of the alpha subunit of eukaryotic initiation factor 2 (eIF2␣) 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 (VV⌬E3L). To further define the underlying mechanism, we first evaluated the effect of pTRS1 on protein kinase R (PKR), the double-stranded RNA (dsRNA)-dependent eIF2␣ kinase. Immunoblot analyses revealed that pTRS1 expression in the context of a VV⌬E3L 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 VV⌬E3L 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 VV⌬E3L replication in HeLa cells. The carboxy terminus of pIRS1 was also required for rescue of VV⌬E3L 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, eIF2␣.Double-stranded RNA (dsRNA) activates the host cell response to viral infection in numerous ways, one of which involves the interferon-induced, dsRNA-dependent protein kinase R (PKR) (reviewed in reference 43). After binding to dsRNA, PKR dimerizes, autophosphorylates, and then phosphorylates the eukaryotic initiation factor 2 (eIF2) on its ␣ subunit. Phosphorylated eIF2␣ sequesters the guanine nucleotide exchange factor eIF2B, resulting in the inhibition of protein synthesis at the level of translation initiation. Since viruses depend on the cellular translational machinery, the shutoff of host cell protein synthesis inhibits viral replication and spread.Many viruses have mechanisms for inhibiting the PKR-mediated phosphorylation of eIF2␣ (56). The vaccinia virus (VV) E3L protein prevents the activation of PKR by binding to and sequestering dsRNA via a carboxy-terminal double-stranded RNA binding domain (dsRBD) (39). VV from which the E3L gene has been deleted (VV⌬E3L) exhibits a dsRNA-dependent restriction of host cell range, and this phenotype can be reversed by expression of the dsRBD from pE3L or dsRNAbinding proteins from other viruses (4, 47, 62). We previously found that the products of the human cytomegalovirus (HCMV) genes TRS1 and IRS1 restore the host cell range of VV⌬E3L, inhibit the phosphorylation of eIF2␣, and prevent the shutoff...

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Section: Discussionmentioning

confidence: 99%

Binding and Nuclear Relocalization of Protein Kinase R by Human Cytomegalovirus TRS1

Hakki

Marshall²,

Niro³

et al. 2006

J Virol

103

100

View full text Add to dashboard Cite

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“…Indirect evidence from use of the PKR inhibitor 2-aminopurine (2-AP) has led to the suggestion that PKR may be acting as a signal transducer involved in the regulation of dsRNA-activated genes. dsRNA treatment of human MG63 osteosarcoma cells induced the expression of the IFN-B gene, which is specifically blocked at the transcriptional level by 2-AP treatment, whereas constitutive -t-actin mRNA levels were unaffected (5). Furthermore, 2-AP has been shown to block the vesicular stomatitis virus induction of IFN in both mouse L cells and chicken embryo cells (6).…”

mentioning

confidence: 99%

Double-stranded RNA-dependent protein kinase activates transcription factor NF-kappa B by phosphorylating I kappa B.

Kumar

Haque

Lacoste

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

518

409

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The induction of interferon (IN) genes by viruses or double-stranded RNA (dsRNA) requires the assembly of a complex set of trnscription factors on responsive DNA elements of ION gene promoters. One of the factors necessary for regulating IFN-P gene transcription is nuclear factor NF-KB, the activation of which is triggered by dsRNA. It has previously been suggested that the dsRNA-activated p68 protein kinase (PKR) may act as an Inducer-receptor, transducing the signal from dsRNA to NF-ucB through phosphorylation of the inhibitor IKB. We present direct evidence that PKR can phosphorylate IKB-a (MAD-3) and activate NF-KB DNA binding activity in vitro. We further show that dsRNA induces an unusual phosphorylated form of IKB-a. The expression of a transdominant mutant PKR is able to perturb the dsRNAmediated signaling pathway in vivo, suggesting a role for this kinase in IFN-P gene induction.

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“…This observation is ascribed to a trans-dominant inhibitory effect of the mutant enzyme on the endogenous wild-type PKR and implicates PKR as a tumor suppressor gene (11)(12)(13)(14). PKR is also implicated in the transcriptional regulation of dsRNA-activated genes, such as interferon ␤, by activation of nuclear transcription factor B (NF B) by phosphorylating its inhibitor, I B, in response to dsRNA (15)(16)(17).…”

mentioning

confidence: 99%

Phosphorylation of Eukaryotic Translation Initiation Factor 2 Mediates Apoptosis in Response to Activation of the Double-stranded RNA-dependent Protein Kinase

Srivastava¹,

Kumar²,

Kaufman³

1998

Journal of Biological Chemistry

358

322

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The interferon-inducible, double-stranded (ds) RNAdependent serine/threonine protein kinase (PKR) plays a role in viral pathogenesis, cell growth, and differentiation and is implicated as a tumor suppressor gene. Expression of a trans-dominant negative, catalytically inactive mutant PKR protected NIH3T3 cells from apoptosis in response to either treatment with tumor necrosis factor ␣ (TNF␣), serum deprivation. In cells expressing mutant PKR, TNF␣, but not dsRNA induced transcription from a nuclear factor B-dependent promoter, demonstrating specificity for dsRNA in signaling through the PKR pathway. Serum or plateletderived growth factor addition to serum-deprived mutant PKR-expressing cells induced transcription of the early response genes c-fos and c-jun, indicating that the immediate early response signaling was intact. Overexpression of wild-type PKR in a transient DNA transfection system was sufficient to induce apoptosis. TNF␣-induced apoptosis correlated with increased phosphorylation of the ␣ subunit of eukaryotic translation initiation factor 2 (eIF-2␣), the primary physiological substrate of the PKR. Furthermore, forced expression of a nonphosphorylatable S51A mutant eIF-2␣ partially protected cells from TNF␣-induced apoptosis, and expression of a S51D mutant eIF-2␣, a mutant that mimics phosphorylated eIF-2␣, was sufficient to induce apoptosis. Taken together, these studies identify a novel requirement for PKR in stress-induced apoptosis that is mediated through eIF-2␣ phosphorylation.

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2-Aminopurine Selectively Inhibits the Induction of β-Interferon, c- fos , and c- myc Gene Expression

Cited by 227 publications

References 19 publications

Binding and Nuclear Relocalization of Protein Kinase R by Human Cytomegalovirus TRS1

Binding and Nuclear Relocalization of Protein Kinase R by Human Cytomegalovirus TRS1

Double-stranded RNA-dependent protein kinase activates transcription factor NF-kappa B by phosphorylating I kappa B.

Phosphorylation of Eukaryotic Translation Initiation Factor 2 Mediates Apoptosis in Response to Activation of the Double-stranded RNA-dependent Protein Kinase

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