Prkra dimer senses double-stranded RNAs to dictate global translation efficiency

Abstract Double-stranded RNAs (dsRNAs), known as conserved pathogen-associated molecular patterns, are recognized by interferon-induced protein kinase R (PKR) to trigger an integrated stress response, characterized by the inhibition of global translation. However, the interferon system is inactive in pluripotent cells, thus the mechanism underlying dsRNA sensing and translational control is unclear. In this study, we utilized early zebrafish embryos as a model of pluripotent cells and discovered a PKR-independent blockage of translation initiation induced by dsRNA stimulation. Prkra dimer was identified as the genuine dsRNA sensor. Upon binding to dsRNAs, the dimerized dsRNA binding domain 3 becomes activated to sequester the eIF2 complexes from the translation machinery, leading to a hindrance in global protein synthesis. This distinctive embryonic stress response restricts RNA virus SVCV replication in zebrafish embryos but is also conserved in early mouse embryos and embryonic stem cells. Therefore, the Prkra-mediated dsRNA sensing and translation blocking mechanism potentially represents a common strategy for reestablishing physiological homeostasis in response to environmental stresses. Competing Interest Statement China patent 202310609693.3 has been authorized to MS, TL and AJC for the application of zebrafish embryos for dsRNA by-product detection. Footnotes In this revision, we elucidated the mode of Prkra activation by demonstrating that it functions as dimers. Upon binding to dsRNA through its first two dsRNA-binding domains (dsRBD1 and dsRBD2), the dimerized dsRBD3 may become activated due to its highly ordered alignment on the dsRNA. The highly ordered binding of Prkra dimer to dsRNA can be visualized in EMSA analysis, manifesting as distinct laddering patterns. We also uncovered a novel mechanism of global translation control, whereby Prkra is activated by dsRNA and utilizes its dimerized dsRBD3 domain to sequester components of the translation initiation complex for the repression of global protein synthesis. This study represents the first report uncovering the functional significance of Prkra dimerization in translational repression. To be more concentrated, we delete the results relating to cell death phenotype in this revised version. In addition, some of the data was revised and errors were corrected. Data availability The deep sequencing data of Ribo-seq are available from NCBI Gene Expression Omnibus GSE234230. The scripts are available at https://github.com/benjaminfang/ribo-seq-workflow. Further information and requests for resources and reagents should be directed to and will be fulfilled by the corresponding author, Ming Shao (shaoming{at}sdu.edu.cn).