Molecular identification and characterization of Heat shock protein 70 family proteins essential for Turnip mosaic virus infection in Arabidopsis thaliana
Abstract
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.