Characterization of Transgenic Mice with Targeted Disruption of the Catalytic Domain of the Double-stranded RNA-dependent Protein Kinase, PKR

Abraham, Ninan; Stojdl, David F.; Duncan, Peter I.; Méthot, Nathalie; Ishii, Tetsu; Dubé, Manon; Vanderhyden, Barbara C.; Atkins, Harold L.; Gray, Douglas A.; McBurney, Michael W.; Koromilas, Antonis E.; Brown, Earl G.; Sonenberg, Nahum; Bell, John C.

doi:10.1074/jbc.274.9.5953

Cited by 216 publications

(187 citation statements)

References 79 publications

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“…Inconsistent with the role of PKR as a tumor suppressor, PKR de®cient or knock-out transgenic mice are normal and do not show any signs of neoplasia (Yang et al, 1995;Abraham et al, 1999). Furthermore, the primary ®broblast cell line derived from the PKR knock-out mouse grows slower than those derived from the normal control mouse (our unpublished observation; Zamanian-Daryoush et al, 1999) implying that PKR may be required for cell proliferation.…”

mentioning

confidence: 53%

Human breast cancer cells contain elevated levels and activity of the protein kinase, PKR

et al. 2000

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mentioning

confidence: 53%

Human breast cancer cells contain elevated levels and activity of the protein kinase, PKR

et al. 2000

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“…13 We examined the biological role of PKR in response to DNA damage in MEFs that are deficient in p53 due to spontaneous immortalization. For this purpose immortalized MEFs completely deficient in PKR activity (PKR À/À MEFs) 14,15 together with their isogenic wild-type counterparts were treated with the chemotherapeutic drug doxorubicin and subjected to analysis of cell death by flow cytometry. We noticed that PKR was required for optimal induction of cell death in response to doxorubicin (Figure 1a).…”

Section: Resultsmentioning

confidence: 99%

“…35 Materials and Methods Cell culture and treatments. PKR À/À MEFs and their isogenic wild-type counterparts 14 were grown in Dulbecco's modified Eagle's medium (DMEM; Wisent, St-Bruno, QC, Canada) supplemented with 10% fetal bovine serum (Wisent) and 100 U/ml of penicillin-streptomycin (Wisent). Isogenic wild-type and PERK À/À MEFs 36 were grown in DMEM supplemented with 10% heat-inactivated bovine serum (Wisent) and 100 U/ml of penicillin-streptomycin.…”

Section: Discussionmentioning

confidence: 99%

Doxorubicin bypasses the cytoprotective effects of eIF2α phosphorylation and promotes PKR-mediated cell death

et al. 2010

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The eukaryotic cell responds to various forms of environmental stress by adjusting the rates of mRNA translation thus facilitating adaptation to the assaulting stress. One of the major pathways that control protein synthesis involves the phosphorylation of the a-subunit of eukaryotic initiation factor eIF2 at serine 51. Different forms of DNA damage were shown to induce eIF2a phosphorylation by using PERK, GCN2 or PKR. However, the specificity of the eIF2a kinases and the biological role of eIF2a phosphorylation pathway in the DNA damage response (DDR) induced by chemotherapeutics are not known. Herein, we show that PKR is the eIF2a kinase that responds to DDR induced by doxorubicin. We show that activation of PKR integrates two signaling pathways with opposing biological outcomes. More specifically, induction of eIF2a phosphorylation has a cytoprotective role, whereas activation of c-jun N-terminal kinase (JNK) by PKR promotes cell death in response to doxorubicin. We further show that the proapoptotic effects of JNK activation prevail over the cytoprotection mediated by eIF2a phosphorylation. These findings reveal that PKR can be an important inducer of cell death in response to chemotherapies through its ability to act independently of eIF2a phosphorylation. The eukaryotic cell has established intricate mechanisms to cope with environmental stress. It does so by changing protein expression in a manner that promotes adaptation to the assaulting stress. Protein expression within the cell is regulated at the transcriptional, translational and posttranslational levels. Translational control is often used under conditions of cellular stress because it allows immediate and selective changes in protein levels. 1 Translation itself is divided into three distinct phases: initiation, elongation and termination. By far the most explored step is that of initiation, which has been shown to be regulated by different extracellular stimuli. 1 One of the major pathways that control translation initiation is that of the phosphorylation of the a-subunit of translation initiation factor eIF2. 2 Phosphorylation of eIF2a at serine 51 (S51) leads to the inhibition of global protein synthesis providing the cell with the opportunity to elicit adaptive responses not only by saving energy but also by preventing the accumulation of unwanted proteins that could interfere with cellular functions. 1 There are four eIF2a kinases that share a homologous kinase domain (KD) but possess different regulatory domains that enable them to become activated by distinct stimuli. 2 The eIF2a kinases are the heme-regulated inhibitor, which is found mainly in erythroid cells and is activated by heme deficiency; the endoplasmic reticulum (ER)-resident kinase (PERK), which is activated by ER stress and inhibits protein synthesis as part of the unfolded protein response; the general aminoacid nonderepressing kinase 2 (GCN2), which responds to amino-acid deprivation and becomes activated by uncharged tRNA and the RNA-dependent protein kinase PKR, an interfer...

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“…Their study assessed inflammasome activity in bone marrow-derived macrophages (BMDMs) isolated from both PKR-ablated mouse strains that were used by either Lu et al [3] or ourselves that targeted either exons 12 [22] or 2-3 [15] of the eIF2αk2 locus, respectively. Although these researchers reported no role for PKR, we contend that close examination of their data from the exon-12-targeted mouse shows that it supports our findings.…”

Section: Discussionmentioning

confidence: 99%

The kinase activity of PKR represses inflammasome activity

Yim

Wang

et al. 2016

Cell Res

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The protein kinase R (PKR) functions in the antiviral response by controlling protein translation and inflammatory cell signaling pathways. We generated a transgenic, knock-in mouse in which the endogenous PKR is expressed with a point mutation that ablates its kinase activity. This novel animal allows us to probe the kinase-dependent and -independent functions of PKR. We used this animal together with a previously generated transgenic mouse that is ablated for PKR expression to determine the role of PKR in regulating the activity of the cryopyrin inflammasome. Our data demonstrate that, in contradiction to earlier reports, PKR represses cryopyrin inflammasome activity. We demonstrate that this control is mediated through the established function of PKR to inhibit protein translation of constituents of the inflammasome to prevent initial priming during innate immune signaling. These findings identify an important role for PKR to dampen inflammation during the innate immune response and caution against the previously proposed therapeutic strategy to inhibit PKR to treat inflammation.

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Characterization of Transgenic Mice with Targeted Disruption of the Catalytic Domain of the Double-stranded RNA-dependent Protein Kinase, PKR

Cited by 216 publications

References 79 publications

Human breast cancer cells contain elevated levels and activity of the protein kinase, PKR

Human breast cancer cells contain elevated levels and activity of the protein kinase, PKR

Doxorubicin bypasses the cytoprotective effects of eIF2α phosphorylation and promotes PKR-mediated cell death

The kinase activity of PKR represses inflammasome activity

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