Zika Virus NS3 Drives the Assembly of a Replication Compartment-like Structure That Exerts the Structural and Physiological Functions of the Viral Replication Compartment

Abstract

Zika virus (ZIKV) is a mosquito-transmitted orthoflavivirus that caused an epidemic in 2015–2016 in the Americas and raised serious global health concerns due to its association with congenital brain anomalies when infections occur during pregnancy. Various viruses can form compartments within the cell to facilitate viral replication and assembly, referred to as viroplasms, replication organelles, or virus factories depending on the type of virus. ZIKV assembles virus particles in virus-generated compartments adjacent to the nucleus, referred to here as a replication compartment (RC), which is formed by remodeling the host cell endoplasmic reticulum (ER). How the viral proteins control RC assembly remains unknown. Here we show that the ZIKV non-structural protein 3 (NS3), a dual-function protease and RNA helicase, is sufficient to drive the assembly of a replication compartment-like structure (RCLS) in human cells. While sufficient to generate the RCLS, NS3 is less efficient in several aspects compared to ZIKV-induced RC assembly. Nonetheless, the RCLS is similar to the ZIKV RC in its assembly at the nuclear periphery, its recruitment of ER, association with the Golgi and centrosome, and the arrangement of microtubules at its surface. Moreover, NS3 expression results in activation of the unfolded protein response (UPR), but attenuates expression of the downstream transcription factor CHOP, mirroring the manipulation of the different aspects of the UPR by ZIKV infection. We further show that the helicase domain and not the protease domain is required for optimal RCLS formation and organelle recruitment, yet each domain affects different control over the UPR. Overall, these findings advance our understanding of the mechanism of RC assembly by ZIKV, its involvement in hijacking the UPR, and the central role of NS3 in the process.