Hijacking a host scaffold protein, RACK1, for replication of a plant RNA virus.

Receptor for activated C kinase 1 (RACK1) is strictly conserved across eukaryotes and acts as a versatile scaffold protein involved in various signaling pathways. Plant RACK1 is known to exert important functions in innate immunity against fungal and bacterial pathogens. However, the role of the RACK1 in plant-virus interactions remains unknown. Here, we addressed the role of RACK1 of Nicotiana benthamiana during infection by red clover necrotic mosaic virus (RCNMV), a plant positive-stranded RNA virus. NbRACK1 was shown to be recruited by the p27 viral replication protein into endoplasmic reticulum-derived aggregated structures (possible replication sites). Downregulation of NbRACK1 by virus-induced gene silencing inhibited viral cap-independent translation and p27-mediated reactive oxygen species (ROS) accumulation, which are prerequisite for RCNMV replication. We also found that NbRACK1 interacted with a host calcium-dependent protein kinase (NbCDPKiso2) that activated a ROS-generating enzyme. Interestingly, NbRACK1 was required for the interaction of p27 with NbCDPKiso2, suggesting that NbRACK1 acts as a bridge between the p27 viral replication protein and NbCDPKiso2. Collectively, our findings provide an example of a viral strategy in which a host multifaceted scaffold protein RACK1 is highjacked for promoting viral protein-triggered ROS production necessary for robust viral replication.