Baculovirus protein PK2 subverts eIF2α kinase function by mimicry of its kinase domain C-lobe.

John J Li, Chune Cao, Sarah M Fixsen, Janet M Young, Chikako Ono, Hisanori Bando, Nels C Elde, Susumu Katsuma, Thomas E Dever, Frank Sicheri

Proceedings of the National Academy of Sciences of the United States of America 2015 Aug 11

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Phosphorylation of eukaryotic translation initiation factor 2α (eIF2α) by eIF2α family kinases is a conserved mechanism to limit protein synthesis under specific stress conditions. The baculovirus-encoded protein PK2 inhibits eIF2α family kinases in vivo, thereby increasing viral fitness. However, the precise mechanism by which PK2 inhibits eIF2α kinase function remains an enigma. Here, we probed the mechanism by which PK2 inhibits the model eIF2α kinase human RNA-dependent protein kinase (PKR) as well as native insect eIF2α kinases. Although PK2 structurally mimics the C-lobe of a protein kinase domain and possesses the required docking infrastructure to bind eIF2α, we show that PK2 directly binds the kinase domain of PKR (PKR(KD)) but not eIF2α. The PKR(KD)-PK2 interaction requires a 22-residue N-terminal extension preceding the globular PK2 body that we term the "eIF2α kinase C-lobe mimic" (EKCM) domain. The functional insufficiency of the N-terminal extension of PK2 implicates a role for the adjacent EKCM domain in binding and inhibiting PKR. Using a genetic screen in yeast, we isolated PK2-activating mutations that cluster to a surface of the EKCM domain that in bona fide protein kinases forms the catalytic cleft through sandwiching interactions with a kinase N-lobe. Interaction assays revealed that PK2 associates with the N- but not the C-lobe of PKR(KD). We propose an inhibitory model whereby PK2 engages the N-lobe of an eIF2α kinase domain to create a nonfunctional pseudokinase domain complex, possibly through a lobe-swapping mechanism. Finally, we show that PK2 enhances baculovirus fitness in insect hosts by targeting the endogenous insect heme-regulated inhibitor (HRI)-like eIF2α kinase.

Citation

John J Li, Chune Cao, Sarah M Fixsen, Janet M Young, Chikako Ono, Hisanori Bando, Nels C Elde, Susumu Katsuma, Thomas E Dever, Frank Sicheri. Baculovirus protein PK2 subverts eIF2α kinase function by mimicry of its kinase domain C-lobe. Proceedings of the National Academy of Sciences of the United States of America. 2015 Aug 11;112(32):E4364-73