YBX1 is required for assembly of viral replication complexes of chikungunya virus and replication of multiple alphaviruses

Chikungunya virus (CHIKV), an enveloped positive-sense RNA virus, is a member of the alphaviruses and cause fever and arthralgia in humans. We performed genome-wide CRISPR/Cas9-based screens and identified Y-box binding protein 1 (YBX1) as an essential cellular factor for CHIKV. Deficiency of YBX1 inhibited CHIKV RNA replication and impaired virus production. Upon CHIKV infection, YBX1 showed a striking re-localization to viral replication complexes (vRCs), where it co-localized with CHIKV nsP3 and dsRNA intermediates. YBX1 directly interacted with CHIKV nsP3, and mutation of the YBX1-binding motif in CHIKV nsP3 suppressed viral replication in host cells. Furthermore, YBX1 bound to viral RNA and increased the viral RNA-binding activity of CHIKV nsP3. Consistently, the RNA-binding activity of YBX1, as well as the ability of nsP3 to bind to YBX1, was required for efficient CHIKV replication. In addition to CHIKV, YBX1 was also essential for replication of all examined alphaviruses including the prototypic alphavirus. Our findings suggest that YBX1 acts as a scaffold for assembly of chikungunya vRCs and an important factor for replication of multiple alphaviruses, which may serve as a potential target for the development of anti-alphavirus therapies.