Yu Huibin, Fernandez Mary Eloise L, Peng Chen, Megawati Dewi, Brennan Greg, Tazi Loubna, Rothenburg Stefan
Department of Medical Microbiology and Immunology, School of Medicine, University of California, Davis, CA 95618, USA.
Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
Vaccines (Basel). 2025 Jul 28;13(8):800. doi: 10.3390/vaccines13080800.
Protein kinase R (PKR) inhibits general mRNA translation by phosphorylating the alpha subunit of eukaryotic translation initiation factor 2 (eIF2). PKR also modulates NF-κB signaling during viral infections, but comparative studies of PKR-mediated NF-κB responses across mammalian species and their regulation by viral inhibitors remain largely unexplored. This study aimed to characterize the conserved antiviral and inflammatory roles of mammalian PKR orthologs and investigate their modulation by poxviral inhibitors. Using reporter gene assays and quantitative RT-PCR, we assessed the impact of 17 mammalian PKR orthologs on general translation inhibition, stress-responsive translation, and NF-κB-dependent induction of target genes. Congenic human and rabbit cell lines infected with a myxoma virus strain lacking PKR inhibitors were used to compare the effects of human and rabbit PKR on viral replication and inflammatory responses. Site-directed mutagenesis was employed to determine key residues responsible for differential sensitivity to the viral inhibitor M156. All 17 mammalian PKR orthologs significantly inhibited general translation, strongly activated stress-responsive ATF4 translation, and robustly induced NF-κB target genes. Inhibition of these responses was specifically mediated by poxviral K3 orthologs that effectively suppressed PKR activation. Comparative analyses showed human and rabbit PKRs similarly inhibited virus replication and induced cytokine transcripts. Amino acid swaps between rabbit PKRs reversed their sensitivity to viral inhibitor M156 and NF-κB activation. Our data show that the tested PKR orthologs exhibit conserved dual antiviral and inflammatory regulatory roles, which can be antagonized by poxviral K3 orthologs that exploit eIF2α mimicry to modulate the PKR-NF-κB axis.
蛋白激酶R（PKR）通过磷酸化真核翻译起始因子2（eIF2）的α亚基来抑制一般的mRNA翻译。PKR在病毒感染期间也调节NF-κB信号传导，但跨哺乳动物物种的PKR介导的NF-κB反应及其受病毒抑制剂调控的比较研究在很大程度上仍未得到探索。本研究旨在表征哺乳动物PKR直系同源物的保守抗病毒和炎症作用，并研究痘病毒抑制剂对它们的调节作用。使用报告基因测定法和定量RT-PCR，我们评估了17种哺乳动物PKR直系同源物对一般翻译抑制、应激反应性翻译以及NF-κB依赖性靶基因诱导的影响。用缺乏PKR抑制剂的黏液瘤病毒株感染同基因的人和兔细胞系，以比较人和兔PKR对病毒复制和炎症反应的影响。采用定点诱变来确定对病毒抑制剂M156差异敏感性的关键残基。所有17种哺乳动物PKR直系同源物均显著抑制一般翻译，强烈激活应激反应性ATF4翻译，并有力地诱导NF-κB靶基因。这些反应的抑制是由痘病毒K3直系同源物特异性介导的，其有效抑制PKR激活。比较分析表明，人和兔PKR同样抑制病毒复制并诱导细胞因子转录本。兔PKR之间的氨基酸交换逆转了它们对病毒抑制剂M156的敏感性和NF-κB激活。我们的数据表明，所测试的PKR直系同源物具有保守的双重抗病毒和炎症调节作用，痘病毒K3直系同源物可通过利用eIF2α模拟来调节PKR-NF-κB轴从而拮抗这些作用。
