Hantaviruses use the endogenous host factor P58IPK to combat the PKR antiviral response

Hantavirus nucleocapsid protein (NP) inhibits protein kinase R (PKR) dimerization by an unknown mechanism to counteract its antiviral responses during virus infection. Here we demonstrate that NP exploits an endogenous PKR inhibitor P58^IPK to inhibit PKR. The activity of P58^IPK is normally restricted in cells by the formation of an inactive complex with its negative regulator Hsp40. On the other hand, PKR remains associated with the 40S ribosomal subunit, a unique strategic location that facilitates its free access to the downstream target eIF2α. Although both NP and Hsp40 bind to P58^IPK, the binding affinity of NP is much stronger compared to Hsp40. P58^IPK harbors an NP binding site, spanning to N-terminal TPR subdomains I and II. The Hsp40 binding site on P58^IPK was mapped to the TPR subdomain II. The high affinity binding of NP to P58^IPK and the overlap between NP and Hsp40 binding sites releases the P58^IPK from its negative regulator by competitive inhibition. The NP-P58^IPK complex is selectively recruited to the 40S ribosomal subunit by direct interaction between NP and the ribosomal protein S19 (RPS19), a structural component of the 40S ribosomal subunit. NP has distinct binding sites for P58^IPK and RPS19, enabling it to serve as bridge between P58^IPK and the 40S ribosomal subunit. NP mutants deficient in binding to either P58^IPK or RPS19 fail to inhibit PKR, demonstrating that selective engagement of P58^IPK to the 40S ribosomal subunit is required for PKR inhibition. Cells deficient in P58^IPK mount a rapid PKR antiviral response and establish an antiviral state, observed by global translational shutdown and rapid decline in viral load. These studies reveal a novel viral strategy in which NP releases P58^IPK from its negative regulator and selectively engages it on the 40S ribosomal subunit to promptly combat the PKR antiviral responses.