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R, K3L or vIF2a, demonstrating that K3 and vIF2a had no effect on yeast cell growth (Figure 2A). In contrast, induction of PKR expression was toxic in the vector-transformed yeast, whereas the toxicity was suppressed by co-expression of K3L or vIF2a (Figure 2B). Based on the homology of vIF2a with eIF2a throughout the entire ORF we tested whether suppression of PKR toxicity might be caused by the
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R, K3L or vIF2a, demonstrating that K3 and vIF2a had no effect on yeast cell growth (Figure 2A). In contrast, induction of PKR expression was toxic in the vector-transformed yeast, whereas the toxicity was suppressed by co-expression of K3L or vIF2a (Figure 2B). Based on the homology of vIF2a with eIF2a throughout the entire ORF we tested whether suppression of PKR toxicity might be caused by the
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Nd zebrafish PKR (Additional file 1: Figure S1B, C). Suppression of PKR toxicity in yeast could be due to impaired PKR expression or due to inhibition of eIF2a phosphorylation. In order to examine eIF2a phosphorylation, yeast whole cell extracts were prepared by the TCA method to prevent protein degradation and dephosphorylation, and Western blot analyses were performed using phospho-specific anti
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Ion of neither vIF2a nor K3L suppressed the growth defects of the sui2-1 mutant strain, we conclude that vIF2a does not functionally substitute for eIF2a. We next compared the effect of vIF2a on human and zebrafish PKR with the effects of the two VACV PKR inhibitors K3 and E3. In the control strain not expressing PKR, expression of K3L or vIF2a had no effect on yeast cell growth, whereas expressio
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Ion of neither vIF2a nor K3L suppressed the growth defects of the sui2-1 mutant strain, we conclude that vIF2a does not functionally substitute for eIF2a. We next compared the effect of vIF2a on human and zebrafish PKR with the effects of the two VACV PKR inhibitors K3 and E3. In the control strain not expressing PKR, expression of K3L or vIF2a had no effect on yeast cell growth, whereas expressio
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The vector pEMBLyex4, were introduced into the temperature-sensitive eIF2a (sui2-1, TD304-10B) mutant strain. The indicated transformants were streaked on SC-Gal medium, where eIF2a expression was maintained and the viral protein expression was induced, and incubated at the indicated temperatures. Results shown are representative of 4 independent transformants for each plasmid.Rothenburg et al. BM
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E 6x coverage mammalian genomes were used to build a profile hidden Markov model with the software HMMER [81], which was then used to screen the platypus ab initio peptide predictions available from Ensembl. An inferred ancestral HIN domain sequence from the node that predates the split between HIN-A, -B and -C domains was also used to perform this search. The genomes of chicken (Gallus gallus rel
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R, K3L or vIF2a, demonstrating that K3 and vIF2a had no effect on yeast cell growth (Figure 2A). In contrast, induction of PKR expression was toxic in the vector-transformed yeast, whereas the toxicity was suppressed by co-expression of K3L or vIF2a (Figure 2B). Based on the homology of vIF2a with eIF2a throughout the entire ORF we tested whether suppression of PKR toxicity might be caused by the