scholarly journals Orthopoxvirus K3 orthologs show virus- and host-specific inhibition of the antiviral protein kinase PKR

2021 ◽  
Vol 17 (1) ◽  
pp. e1009183
Author(s):  
Chorong Park ◽  
Chen Peng ◽  
M. Julhasur Rahman ◽  
Sherry L. Haller ◽  
Loubna Tazi ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Vaccinia Virus ◽  
Host Range ◽  
Translation Initiation Factor ◽  
Host Cells ◽  
Initiation Factor ◽  
Specific Inhibition ◽  
Antiviral Protein ◽  
Range Function ◽  
Species Specific

The antiviral protein kinase R (PKR) is an important host restriction factor, which poxviruses must overcome to productively infect host cells. To inhibit PKR, many poxviruses encode a pseudosubstrate mimic of the alpha subunit of eukaryotic translation initiation factor 2 (eIF2), designated K3 in vaccinia virus. Although the interaction between PKR and eIF2α is highly conserved, some K3 orthologs from host-restricted poxviruses were previously shown to inhibit PKR in a species-specific manner. To better define this host range function, we compared the sensitivity of PKR from 17 mammals to inhibition by K3 orthologs from closely related orthopoxviruses, a genus with a generally broader host range. The K3 orthologs showed species-specific inhibition of PKR and exhibited three distinct inhibition profiles. In some cases, PKR from closely related species showed dramatic differences in their sensitivity to K3 orthologs. Vaccinia virus expressing the camelpox virus K3 ortholog replicated more than three orders of magnitude better in human and sheep cells than a virus expressing vaccinia virus K3, but both viruses replicated comparably well in cow cells. Strikingly, in site-directed mutagenesis experiments between the variola virus and camelpox virus K3 orthologs, we found that different amino acid combinations were necessary to mediate improved or diminished inhibition of PKR derived from different host species. Because there is likely a limited number of possible variations in PKR that affect K3-interactions but still maintain PKR/eIF2α interactions, it is possible that by chance PKR from some potential new hosts may be susceptible to K3-mediated inhibition from a virus it has never previously encountered. We conclude that neither the sensitivity of host proteins to virus inhibition nor the effectiveness of viral immune antagonists can be inferred from their phylogenetic relatedness but must be experimentally determined.

scholarly journals Orthopoxvirus K3 orthologs show virus- and host-specific inhibition of the antiviral protein kinase PKR

2020 ◽  
Author(s):  
Chorong Park ◽  
Chen Peng ◽  
M. Julhasur Rahman ◽  
Sherry L. Haller ◽  
Loubna Tazi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Protein Kinase ◽  
Vaccinia Virus ◽  
Related Species ◽  
Translation Initiation Factor ◽  
Virus Species ◽  
Specific Inhibition ◽  
Antiviral Protein ◽  
Closely Related Species ◽  
Species Specific

AbstractThe antiviral protein kinase R (PKR) is an important host restriction factor, which poxviruses must overcome to productively infect host cells. To inhibit PKR, many poxviruses encode a pseudosubstrate mimic of the alpha subunit of eukaryotic translation initiation factor 2 (eIF2), designated K3 in vaccinia virus. Although the interaction between PKR and eIF2α is highly conserved, some K3 orthologs were previously shown to inhibit PKR in a species-specific manner. To better define this host range function, we compared the sensitivity of PKR from 17 mammals to inhibition by K3 orthologs from closely related orthopoxviruses. The K3 orthologs showed species-specific inhibition of PKR and exhibited three distinct inhibition profiles. In some cases, PKR from closely related species showed dramatic differences in their sensitivity to K3 orthologs. Vaccinia virus expressing the camelpox virus K3 ortholog replicated more than three orders of magnitude better in human and sheep cells than a virus expressing vaccinia virus K3, but both viruses replicated comparably well in cow cells. Strikingly, in site-directed mutagenesis experiments between the variola virus and camelpox virus K3 orthologs, we found that different amino acid combinations were necessary to mediate improved or diminished inhibition of PKR derived from different host species. Because there is likely a limited number of possible variations in PKR that affect K3-interactions but still maintain PKR/eIF2α interactions, by chance PKR from some potential new hosts may be susceptible to K3-mediated inhibition from a virus it has never previously encountered. We conclude that neither the sensitivity of host proteins to virus inhibition nor the effectiveness of viral immune antagonists can be inferred from their phylogenetic relatedness but must be experimentally determined.Authors summaryMost virus families are composed of large numbers of virus species. However, in general, only a few prototypic viruses are experimentally studied in-depth, and it is often assumed that the obtained results are representative of other viruses in the same family. In order to test this assumption, we compared the sensitivity of the antiviral protein kinase PKR from various mammals to inhibition by multiple orthologs of K3, a PKR inhibitor expressed by several closely related orthopoxviruses. We found strong differences in PKR inhibition by the K3 orthologs, demonstrating that sensitivity to a specific inhibitor was not indicative of broad sensitivity to orthologs of these inhibitors from closely related viruses. We also show that PKR from even closely related species displayed markedly different sensitivities to these poxvirus inhibitors. Furthermore, we identified amino acid residues in these K3 orthologs that are critical for enhanced or decreased PKR inhibition and found that distinct amino acid combinations affected PKRs from various species differently. Our study shows that even closely related inhibitors of an antiviral protein can vary dramatically in their inhibitory potential, and cautions that results from host-virus interaction studies of a prototypic virus genus member cannot necessarily be extrapolated to other viruses in the same genus without experimental verification.

scholarly journals Species-specific inhibition of capripoxvirus replication by host antiviral protein kinase R

2018 ◽  
Vol 1438 (1) ◽  
pp. 3-17 ◽  
Author(s):  
Zhixun Zhao ◽  
Xueliang Zhu ◽  
Na Wu ◽  
Xiaodong Qin ◽  
Caiyun Huang ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Specific Inhibition ◽  
Antiviral Protein ◽  
Protein Kinase R ◽  
Species Specific

scholarly journals Vaccinia Virus K1L and C7L Inhibit Antiviral Activities Induced by Type I Interferons

2009 ◽  
Vol 83 (20) ◽  
pp. 10627-10636 ◽  
Author(s):  
Xiangzhi Meng ◽  
Canhua Jiang ◽  
Janilyn Arsenio ◽  
Kevin Dick ◽  
Jingxin Cao ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Host Range ◽  
Mammalian Cells ◽  
Initiation Factor ◽  
Viral Gene ◽  
Type I ◽  
Type I Ifn ◽  
Α Subunit ◽  
Range Function ◽  
Huh7 Cells

ABSTRACT Cellular tropism of vaccinia virus (VACV) is regulated by host range genes, including K1L, C7L, and E3L. While E3L is known to support viral replication by antagonizing interferon (IFN) effectors, including PKR, the exact functions of K1L and C7L are unclear. Here, we show that K1L and C7L can also inhibit antiviral effectors induced by type I IFN. In human Huh7 and MCF-7 cells, a VACV mutant lacking both K1L and C7L (vK1L−C7L−) replicated as efficiently as wild-type (WT) VACV, even in the presence of IFN. However, pretreating the cells with type I IFN, while having very little effect on WT VACV, blocked the replication of vK1L−C7L− at the step of intermediate viral gene translation. Restoring either K1L or C7L to vK1L−C7L− fully restored the IFN resistance phenotype. The deletion of K1L and C7L from VACV did not affect the ability of the virus to inhibit IFN signaling or its ability to inhibit the phosphorylation of PKR and the α subunit of eukaryotic initiation factor 2, indicating that K1L and C7L function by antagonizing an IFN effector(s) but with a mechanism that is different from those of IFN antagonists previously identified for VACV. Mutations of K1L that inactivate the host range function also rendered K1L unable to antagonize IFN, suggesting that K1L supports VACV replication in mammalian cells by antagonizing the same antiviral factor(s) that is induced by IFN in Huh7 cells.

scholarly journals Palmitate Causes Endoplasmic Reticulum Stress and Apoptosis in Human Mesenchymal Stem Cells: Prevention by AMPK Activator

Endocrinology ◽  
2012 ◽  
Vol 153 (11) ◽  
pp. 5275-5284 ◽  
Author(s):  
Jun Lu ◽  
Qinghua Wang ◽  
Lianghu Huang ◽  
Huiyue Dong ◽  
Lingjing Lin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Endoplasmic Reticulum ◽  
Mesenchymal Stem Cells ◽  
Protein Kinase ◽  
Er Stress ◽  
Human Mesenchymal Stem Cells ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Human Umbilical Cord ◽  
Amp Activated Protein Kinase

Abstract Elevated circulating saturated fatty acids concentration is commonly associated with poorly controlled diabetes. The highly prevalent free fatty acid palmitate could induce apoptosis in various cell types, but little is known about its effects on human mesenchymal stem cells (MSCs). Here, we report that prolonged exposure to palmitate induces human bone marrow-derived MSC (hBM-MSC) and human umbilical cord-derived MSC apoptosis. We investigated the role of endoplasmic reticulum (ER) stress, which is known to promote cell apoptosis. Palmitate activated XBP1 splicing, elF2α (eukaryotic translation initiation factor 2α) phosphorylation, and CHOP, ATF4, BiP, and GRP94 transcription in hBM-MSCs. ERK1/2 and p38 MAPK phosphorylation were also induced by palmitate in hBM-MSCs. A selective p38 inhibitor inhibited palmitate activation of the ER stress, whereas the ERK1/2 inhibitors had no effect. The AMP-activated protein kinase activator aminoimidazole carboxamide ribonucleotide blocked palmitate-induced ER stress and apoptosis. These findings suggest that palmitate induces ER stress and ERK1/2 and p38 activation in hBM-MSCs, and AMP-activated protein kinase activator prevents the deleterious effects of palmitate by inhibiting ER stress and apoptosis.

Translation of the yeast transcriptional activator GCN4 is stimulated by purine limitation: implications for activation of the protein kinase GCN2

1993 ◽  
Vol 13 (8) ◽  
pp. 5099-5111
Author(s):  
R J Rolfes ◽  
A G Hinnebusch
Keyword(s):  
Amino Acid ◽  
Protein Kinase ◽  
Transcriptional Activation ◽  
Phosphorylation Site ◽  
Transcriptional Activator ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Single Amino Acid ◽  
Biosynthetic Genes ◽  
Transcriptional Activator Protein

The transcriptional activator protein GCN4 is responsible for increased transcription of more than 30 different amino acid biosynthetic genes in response to starvation for a single amino acid. This induction depends on increased expression of GCN4 at the translational level. We show that starvation for purines also stimulates GCN4 translation by the same mechanism that operates in amino acid-starved cells, being dependent on short upstream open reading frames in the GCN4 mRNA leader, the phosphorylation site in the alpha subunit of eukaryotic translation initiation factor 2 (eIF-2 alpha), the protein kinase GCN2, and translational activators of GCN4 encoded by GCN1 and GCN3. Biochemical experiments show that eIF-2 alpha is phosphorylated in response to purine starvation and that this reaction is completely dependent on GCN2. As expected, derepression of GCN4 in purine-starved cells leads to a substantial increase in HIS4 expression, one of the targets of GCN4 transcriptional activation. gcn mutants that are defective for derepression of amino acid biosynthetic enzymes also exhibit sensitivity to inhibitors of purine biosynthesis, suggesting that derepression of GCN4 is required for maximal expression of one or more purine biosynthetic genes under conditions of purine limitation. Analysis of mRNAs produced from the ADE4, ADE5,7, ADE8, and ADE1 genes indicates that GCN4 stimulates the expression of these genes under conditions of histidine starvation, and it appeared that ADE8 mRNA was also derepressed by GCN4 in purine-starved cells. Our results indicate that the general control response is more global than was previously imagined in terms of the type of nutrient starvation that elicits derepression of GCN4 as well as the range of target genes that depend on GCN4 for transcriptional activation.

scholarly journals The Potential Role of Protein Kinase R as a Regulator of Age-Related Neurodegeneration

2021 ◽  
Vol 13 ◽  
Author(s):  
Nicolás W. Martinez ◽  
Felipe E. Gómez ◽  
Soledad Matus
Keyword(s):  
Protein Kinase ◽  
Pathological Process ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Protein Kinase R ◽  
Eukaryotic Translation ◽  
Neurodegenerative Process ◽  
Age Related ◽  
Initiation Factor 2

There is a growing evidence describing a decline in adaptive homeostasis in aging-related diseases affecting the central nervous system (CNS), many of which are characterized by the appearance of non-native protein aggregates. One signaling pathway that allows cell adaptation is the integrated stress response (ISR), which senses stress stimuli through four kinases. ISR activation promotes translational arrest through the phosphorylation of the eukaryotic translation initiation factor 2 alpha (eIF2α) and the induction of a gene expression program to restore cellular homeostasis. However, depending on the stimulus, ISR can also induce cell death. One of the ISR sensors is the double-stranded RNA-dependent protein kinase [protein kinase R (PKR)], initially described as a viral infection sensor, and now a growing evidence supports a role for PKR on CNS physiology. PKR has been largely involved in the Alzheimer’s disease (AD) pathological process. Here, we reviewed the antecedents supporting the role of PKR on the efficiency of synaptic transmission and cognition. Then, we review PKR’s contribution to AD and discuss the possible participation of PKR as a player in the neurodegenerative process involved in aging-related pathologies affecting the CNS.

scholarly journals A Poxvirus Host Range Protein, CP77, Binds to a Cellular Protein, HMG20A, and Regulates Its Dissociation from the Vaccinia Virus Genome in CHO-K1 Cells

2006 ◽  
Vol 80 (15) ◽  
pp. 7714-7728 ◽  
Author(s):  
Jye-Chian Hsiao ◽  
Chien-Chiang Chao ◽  
Ming-Jer Young ◽  
Yu-Tai Chang ◽  
Er-Chieh Cho ◽  
...  
Keyword(s):  
Amino Acids ◽  
Vaccinia Virus ◽  
Host Range ◽  
Viral Genome ◽  
Cellular Protein ◽  
Deletion Mapping ◽  
Viral Dna ◽  
Virus Growth ◽  
Range Function ◽  
Viral Host Range

ABSTRACT Vaccinia virus does not grow in Chinese hamster ovary (CHO-K1) cells in the absence of a viral host range factor, cowpox protein CP77. In this study, CP77 was fused to the C terminus of green fluorescence protein (GFP-CP77) and a series of nested deletion mutants of GFP-CP77 was constructed for insertion into a vaccinia virus host range mutant, VV-hr, and expressed from a viral early promoter. Deletion mapping analyses demonstrated that the N-terminal 352 amino acids of CP77 were sufficient to support vaccinia virus growth in CHO-K1 cells, whereas the C-terminal residues 353 to 668 were dispensable. In yeast two-hybrid analyses, CP77 bound to a cellular protein, HMG20A, and GST pulldown analyses showed that residues 1 to 234 of CP77 were sufficient for this interaction. After VV-hr virus infection of CHO-K1 cells, HMG20A was translocated from the nucleus to viral factories and bound to the viral genome via the HMG box region. In control VV-hr-infected CHO-K1 cells, binding of HMG20A to the viral genome persisted from 2 to 8 h postinfection (h p.i.); in contrast, when CP77 was expressed, the association of HMG20A with viral genome was transient, with little HMG20A remaining bound at 8 h p.i. This indicates that dissociation of HMG20A from viral factories correlates well with CP77 host range activity in CHO-K1 cells. Finally, in cells expressing a CP77 deletion protein (amino acids 277 to 668) or a ΔANK5 mutant that did not support vaccinia virus growth and did not contain the HMG20A binding site, HMG20A remained bound to viral DNA, demonstrating that the binding of CP77 to HMG20A is essential for its host range function. In summary, our data revealed that a novel cellular protein, HMG20A, the dissociation of which from viral DNA is regulated by CP77, providing the first cellular target regulated by viral host range CP77 protein.

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