Contribution of dsRBD2 to PKR Activation

Protein kinase R (PKR) is a key pattern recognition receptor of the innate immune pathway. PKR is activated by double-stranded RNA (dsRNA) that is often produced during viral genome replication and transcription. PKR contains two tandem double-stranded RNA binding domains at the N-terminus, dsRBD1 and dsRBD2, and a C-terminal kinase domain. In the canonical model for activation, RNAs that bind multiple PKRs induce dimerization of the kinase domain that promotes an active conformation. However, there is evidence that dimerization of the kinase domain is not sufficient to mediate activation and PKR activation is modulated by the RNA-binding mode. dsRBD2 lacks most of the consensus RNA-binding residues, and it has been suggested to function as a modulator of PKR activation. Here, we demonstrate that dsRBD2 regulates PKR activation and identify the N-terminal helix as a critical region for modulating kinase activity. Mutations in dsRBD2 that have minor effects on overall dsRNA-binding affinity strongly inhibit the activation of PKR by dsRNA. These mutations also inhibit RNA-independent PKR activation. These data support a model where dsRBD2 has evolved to function as a regulator of the kinase.