Wu, Guanwei, Jia, Zhaoxing, Ding, Kaida, Zheng, Hongying, Lu, Yuwen, Lin, Lin, Peng, Jiejun, Rao, Shaofei, Wang, Aiming, Chen, Jianping, and Yan, Fei
Accumulated experimental evidence has shown that viruses recruit the host intracellular machinery to establish infection. It has recently been shown that the potyvirus Turnip mosaic virus (TuMV) transits through the late endosome (LE) for viral genome replication, but it is still largely unknown how the viral replication vesicles labelled by the TuMV membrane protein 6K2 target LE. To further understand the underlying mechanism, we studied the involvement of the vacuolar sorting receptor (VSR) family proteins from Arabidopsis in this process. We now report the identification of VSR4 as a new host factor required for TuMV infection. VSR4 interacted specifically with TuMV 6K2 and was required for targeting of 6K2 to enlarged LE. Following overexpression of VSR4 or its recycling-defective mutant that accumulates in the early endosome (EE), 6K2 did not employ the conventional VSR-mediated EE to LE pathway, but targeted enlarged LE directly from cis-Golgi and viral replication was enhanced. In addition, VSR4 can be N-glycosylated and this is required for its stability and for monitoring 6K2 trafficking to enlarged LE. A non-glycosylated VSR4 mutant enhanced the dissociation of 6K2 from cis-Golgi, leading to the formation of punctate bodies that targeted enlarged LE and to more robust viral replication than with glycosylated VSR4. Finally, TuMV hijacks N-glycosylated VSR4 and protects VSR4 from degradation via the autophagy pathway to assist infection. Taken together, our results have identified a host factor VSR4 required for viral replication vesicles to target endosomes for optimal viral infection and shed new light on the role of N-glycosylation of a host factor in regulating viral infection. Author summary: A key feature of the replication of positive-strand RNA viruses is the rearrangement of the host endomembrane system to produce a membranous replication organelle. Recent reports suggest that the late endosome (LE) serves as a replication site for the potyvirus Turnip mosaic virus (TuMV), but the mechanism(s) by which TuMV replication vesicles target LE are far from being fully elucidated. Identification of the host factors involved in this transport process could lead to new strategies to combat TuMV infection. In this report, we provide evidence that TuMV replication depends on functional vesicle transport from cis-Golgi to the enlarged LE pathway that is mediated by a specific VSR family member, VSR4, from Arabidopsis. Knock out of VSR4 impaired the targeting of TuMV replication vesicles to enlarged LE and suppressed viral infection, and this process depends on the specific interaction between VSR4 and the viral replication vesicle-forming protein 6K2. We also showed that N-glycosylation of VSR4 modulates the targeting of TuMV replication vesicles to enlarged LE and enhances viral infection, thus contributing to our understanding of how TuMV manipulates host factors in order to establish optimal infection. These results may have implications for the role of VSR in other positive-strand RNA viruses. [ABSTRACT FROM AUTHOR]