Abstract
Herpesvirus genomes are decoded by host RNA polymerase enzymes, generating mRNAs that are post-transcriptionally modified and exported to the cytoplasm through the combined work of host and viral factors. These viral mRNAs bear 5′-m7G caps and poly(A) tails that should permit assembly of canonical eIF4F cap-binding complexes to initiate protein synthesis. However, the precise mechanisms of translation initiation remain to be investigated for most herpesviruses. eIF4F assembly requires mTORC1-dependent phosphorylation of 4E-BP1, which releases eIF4E from repressive protein complexes. Here, we report that mTORC1 is active and eIF4F is readily detectable throughout the Kaposi’s sarcoma-associated herpesvirus (KSHV) replication cycle. Pharmacologic inhibition of mTORC1 activity caused eIF4F disassembly in KSHV-infected cells, indicating that the mTORC1/4E-BP1/eIF4F signalling axis was intact during virus replication. mTORC1 activity contributed to global protein synthesis in infected cells and was required for lytic gene expression immediately upon reactivation from latency. However, once early gene expression had begun, mTORC1 activity was largely dispensable for viral genome replication, late gene expression, and release of infectious viral progeny. Furthermore, polysome fractionation and RNA-sequencing analysis demonstrated that the translational efficiency of viral mRNA was unaltered by changes in the abundance of eIF4F. Accumulating evidence suggests that herpesvirus mRNA translation can be initiated in an eIF4F-independent manner by an alternative mTORC1/4EBP1-resistant initiation complex, thereby facilitating translation of viral mRNA and the production of infectious progeny.
Author summary All viruses require complex host cell machinery to convert viral mRNAs into proteins. Efficient protein synthesis is required to meet the high anabolic demands of a growing cell. A protein complex known as mechanistic target of rapamycin complex 1 (mTORC1) is a master regulator of protein synthesis. In nutrient-rich conditions, mTORC1 is active and promotes protein synthesis. In nutrient-poor conditions or under stress, mTORC1 is rapidly shut off, and global protein synthesis is arrested. Remarkably, we discovered that synthesis of Kaposi’s sarcoma-associated herpesvirus (KSHV) proteins is largely resistant to drugs that inhibit mTORC1, including rapamycin and a more recently developed drug known as Torin. These surprising findings suggest that herpesviruses may employ previously unknown mechanisms to ensure efficient synthesis of viral proteins in situations when canonical translation initiation machinery would be expected to succumb to stress.