Electronic Thesis and Dissertation Repository

Thesis Format



Doctor of Philosophy




Wang, Aiming.


London Research and Development Centre, Agriculture and Agri-Food Canada (AAFC)

2nd Supervisor

Bernards, Mark A.



Turnip mosaic virus (TuMV) belongs to the RNA virus family of Potyviridae and genus Potyvirus. TuMV incurs agricultural losses by causing diseases in vegetable, oilseed, forage, and biofuel crops globally. Viruses are obligate intracellular parasites depending on the host cellular machinery to proliferate. Thus, molecular identification and functional characterization of host factors essential in the viral infection process may open up a new avenue towards developing genetic virus resistance. Eukaryotic translation initiation factor 4E (eIF4E) or its isoform (eIF(iso)4E) is a critical host factor for many potyviruses including TuMV. Heat shock protein 70 family proteins (HSP70) have been identified in the eIF(iso)4E protein complex isolated from Arabidopsis thaliana infected with TuMV. I hypothesized that at least some A. thaliana HSP70s are host factors for TuMV infection since they are associated with the most essential and multiple functional TuMV host factor eIF(iso)4E. To explore the roles of HSP70s in TuMV infection, TuMV infection assay in A. thaliana HSP70 mutants were performed. Combining the HSP70s identified from eIF(iso)4E protein complex, five HSP70s: cytoplasmic HSP70-1, HSP70-2, and HSP70-8, as well as endoplasmic reticulum (ER)- located HSP70-11 and HSP70-12 were selected for further analysis of their involvement in TuMV infection using different systems. I confirmed interaction of all five HSP70s with eIF(iso)4E and discovered their interactions with TuMV viral proteins: replicase NIb and coat protein CP. I found that the HSP70s colocalized with TuMV replication complex in infected plant leaf cells. HSP70-1 and HSP70-2 have mostly inhibitory effect on TuMV infection by accelerating the degradation of NIb via the ubiquitin-proteosome pathway. Consistent to my hypothesis, HSP70-11 and HSP70-12 facilitate TuMV infection, probably by inhibiting the degradation of TuMV protein NIb and CP as well as the host factor eIF(iso)4E. The proviral effect of HSP70-11 and HSP70-12 possibly depends on their ER-localization. Finally, the effect of HSP70-8 on TuMV infection was ambiguous. While HSP70-8 accelerated the degradation of TuMV NIb and CP, TuMV infection was reduced in HSP70-8 knockout A. thaliana plants. Together, these data suggest that HSP70s play complex roles in TuMV, possibly associated with their chaperone activities on different viral proteins and host factors.

Summary for Lay Audience

Viruses rely on invading host cells for survival and multiplication because they are too simple to produce their own material and energy. They need to use the proteins from the host cell to survive and reproduce. These host proteins are named host factors. When viruses infect hosts, they may cause diseases or even death. Studying host factors increases knowledge on how viruses infect hosts and how to control diseases caused by viruses. One way to stop virus from infecting a host is to disable one or more host factors, preventing the virus from using them. This is a useful way to fight virus infection especially in plants, as plants usually have more than one version of proteins with the same function. Therefore, disabling one essential host factor does not harm the normal growth of the plant because it still has other versions to use. In this thesis, I studied Turnip mosaic virus (TuMV), which causes problems in many important crops. We looked into a family of host proteins called the Heat shock protein 70s (HSP70) to see if TuMV takes advantage of them as host factors for infection. In plants, HSP70s are responsible for maintaining the protein’s proper shape and location and balancing cellular protein production and recycling. We found that two HSP70s located in the endoplasmic reticulum (ER), or the membrane system within the cell, are host factors for TuMV infection. They are found in the same place where the virus replicates within the host, and they interact with virus protein responsible for virus replication. They may help virus infection by preventing the virus protein from breaking down. On the other hand, the HSP70s located in the cytoplasm have an opposite effect on TuMV infection compared with two HSP70s located in the ER. Though they also locate in the virus replication compartments and interact with virus protein, the cytoplasm HSP70s have a negative impact on TuMV infection by promoting the degradation of virus protein. Moreover, if the ER-located HSP70s were modified so that they did not target the ER, the modified HSP70s acted like the cytoplasm-located HSP70s and inhibited TuMV infection.