DsRNA-dependent protein kinase

Gil, J., and M. Esteban. 2000. Induction of apoptosis by the dsRNA-dependent protein kinase (PKR) Mechanism of action. Apoptosis 5 107-14. 

This is converted to an inactive phosphorylated form by a dsRNA-dependent protein kinase (Fig. 31-10). The protein kinase also appears to be an interferon-induced protein as is the oligo(2 -5 A)-activated RNAse indicated in Fig. 31-10. ° Interferons have effects other than inducing the antiviral state. Thus, human IFN-p2 is identical to a B-cell differentiation factor. Both IFN-a and IFN-p have antigrowth activity and are currently in use for treatment of some forms of cancer as well as for viral infections. ... 

Crude lysates (S10) or ribosomal extracts from L cell cultures treated for 24 h by 200 U/ml mouse interferon, when incubated with (32p) y (pp low concentrations of dsRNA (O.O4 - 0.4 M-g/nil poly rI rC), demonstrate an increased phosphorylation of several endogenous proteins (4"9) Among these, proteins 67 and 55 (67,000 and 55,000 MW in SDS) are most prominent. Protein 55 comigrates by polyacrylamide gel electrophoresis with the small subunit of initiation factor eIF-2 (Figure 2). Interferon-treated cell extracts contain a dsRNA-dependent protein kinase phosphorylating this subunit of eIF-2, this activity being more... 

It is, therefore, likely, that phosphorylation of eIF-2 impairs one of its function required for met-tRNA binding to 4OS ribosomes, as shown recently in reticulocyte lysate for the haemin-regulated eIF-2 phosphorylation (14, I5). A role for the other ribosome-associated proteins phosphorylated by the dsRNA-dependent protein kinase, e.g. protein 67 and I7 (4)> "the translational inhibition is, however, not excluded. Factor Ml may also be involved (5) ... 

R 291 R.P. Barnwal, S. Girdhani and S. Nanduri, NMR Studies towards Understanding the Activation Mechanism of dsRNA Dependent Protein Kinase, PKR , P.Ind.Natl.Sci.Acad., A, Phys.Sci., 2004,70,597... 

Wu, S. and R.J. Kaufman, Double-stranded (ds) RNA binding and not dimerization correlates with the activation of the dsRNA-dependent protein kinase (PKR). J Biol Chem, 1996.271(3) p. 1756-63. 

H. et al.. Mouse 3-phosphoinositide-dependent protein kinase-1 undergoes dimerization and franx-phosphorylation in the activation loop, J. Biol. Chem. 278, 42913 2919, 2003 Wu, S. and Kaufman, R.J., trans-auto-phosphorylation by the isolated kinase domain is not sufQcient for dimerization or activation of the dsRNA-activated protein kinase PKR, Biochemistry 43, 11027-11034, 2004. 

Double-stranded (ds) RNA-dependent protein kinase (PKR) was the first PRR shown to rect ise a product of viral replication, namely dsRNA. PKR is induced by IFN and in a critical host antiviral defence mechanism and as such it is both a PRR and an IFN eflFeaor protein. The discovery of PKR explained the observation that in cells pretreated with IFN, the translation ofviral mRNAs were blocked. - PKR is activated in response to viral, cellular or synthetic dsRNA of 30... 

During purification of the protein synthesis inhibitors from interferon-treated cells, the dsENA-ATP activated protein kinase PK-i, was separated from another dsRNA-ATP dependent activity. 

In mouse L cells, addition of one or two minor species of mammalian leucyl-tRNA were shown to be sufficient to restore translation (27). A similar conclusion was reached with yeast tRNA (28), even with poly(U,C) as template. The type of tRNA to be added appears to depend on the mRNA used (25, 27) and also on the extract Friend cells were reported to require lysyl-tRNA (29). These tRNA species were, however, not absent from interferon-treated cell extracts (24, 27, 50), l> u.t had to be added in excess over what is normally needed. A decreased charging of leu-tRNA or an increased deacylation were reported (5I, 32). Prolonged pre-incubation for 1-2 h of the extracts can increase the interferon-induced elongation block reversed by tRNA (55)-have recently observed that ATP is required to activate the translational inhibition seen in interferon-treated cell extracts without dsRNA. This could be shown by comparing extracts freed of endogenous mRNA activity either by micrococcal nuclease (54) or by pre-incubation with ATP. Table 5 shows that tRNA reversible-inhibition developed when ATP was present. It is not clear yet whether the protein kinase, or E and F may play a role in this inhibition without dsRNA. We also observed an inhibitor of leu-tRNA charging in ribosomal extracts of interferon-treated cells (Schmidt, A. and Revel, M., impublished). 

These observations can account for the greater sensitivity of translation to dsRNA in extracts from interferon-treated cells. Apparently, interferon treatment induces a rise in the level of dsRNA-dependent eIF-2-kinase, but since the enzyme is inactive in the absence of sufficient dsRNA, protein synthesis in interferon treated, uninfected cells continues normally. During infection, however, virus-generated dsRNA potentiates the kinase and the resulting extensive phosphorylation of eIF-2 leads to a general inhibition of initiation of translation. As noted for reticulocyte lysates, in extracts of interferon-treated cells, too, there is no good correlation between the extent of phosphorylation of eIF-2 and inhibition of translation (Jacobsen et al., 1983). 

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