PKR is the RNA-activated protein kinase and mediates an anti-viral response in humans. It is activated by long stretches of perfect dsRNA to autophosphorylate and then phosphorylate eIF-2α, thereby inhibiting the initiation of translation. It has a dsRNA binding domain (dsRBD) that consists of two dsRBMs. We showed that PKR has a site size of about 16 bp and that it is activated by a variety of non-canonical RNA structures. For instance, PKR can be activated by short dsRNAs, as long as they have single-stranded tails. Our lab demonstrated that the 5′-triphosphate acts as a pathogen associated molecular patter (PAMP) to activate PKR and initiate translation inhibition. Additional studies showed roles for RNA dimerization and PKR dimerization and activation, and roles for RNA modifications and bulges in PKR regulation.
One recent study from our lab demonstrated that native tertiary structure and nucleoside modifications suppress tRNA’s intrinsic ability to activate PKR . We also showed that PKR can straighten bent dsRNAs using both gel electrophoresis and SAXS methods  and that a variety of misfolded RNAs can activate PKR including tRNA dimers and misfolded versions of certain ribozymes . These findings have implications for the ability of dsRMBs to remodel RNA structure, and for roles of RNA folding in disease. We are currently working to understand how RNAs from new and novel pathogens activate PKR both in the cell and under in vivo-like conditions in the lab. Both molecular biology and biophysical approaches are used.
Kinetics, mutagenesis, RNA-protein interactions, ITC, SAXS, CLIP
1. Nallagatla, S. R., Hwang, J., Toroney, R., Zheng, X., Cameron, C. E. & Bevilacqua, P. C. (2007). 5’-triphosphate-dependent activation of PKR by RNAs with short stem-loops. Science 318, 1455-1458.
2. Patel, S., Blose, J. M., Sokoloski, J. E., Pollack, L. & Bevilacqua, P. C. (2012). Specificity of the double-stranded RNA-binding domain from the RNA-activated protein kinase PKR for double-stranded RNA: insights from thermodynamics and small-angle X-ray scattering. Biochemistry 51, 9312-9322.
3. Heinicke, L. A. & Bevilacqua, P. C. (2012). Activation of PKR by RNA misfolding: HDV ribozyme dimers activate PKR. RNA 18, 2157-2165.
4. Toroney, R., Nallagatla, S. R., Boyer, J. A., Cameron, C. E. & Bevilacqua, P. C. (2010). Regulation of PKR by HCV IRES RNA: importance of domain II and NS5A. J. Mol. Biol. 400, 393-412.
5. Toroney, R., Hull, C. M., Sokoloski, J. E. & Bevilacqua, P. C. (2012). Mechanistic characterization of the 5′-triphosphate-dependent activation of PKR: Lack of 5′-end nucleobase specificity, evidence for a distinct triphosphate binding site, and a critical role for the dsRBD. RNA 18, 1862-1874.
1. Nallagatla, S. R., Toroney, R. & Bevilacqua, P. C. (2008). A brilliant disguise for self RNA: 5′-end and internal modifications of primary transcripts suppress elements of innate immunity. RNA Biol. 5, 140-144. [pdf]
2. Nallagatla, S. R., Toroney, R. & Bevilacqua, P. C. (2011). Regulation of innate immunity through RNA structure and the protein kinase PKR. Curr. Opin. Struct. Biol. 21, 119-127.[pdf]