Characterization of Japanese soil-borne wheat mosaic virus movement protein and investigation of interacting host-plant factors

Abstract

The Japanese soil-borne wheat mosaic virus (JSBWMV) is a cereal virus infecting barley as well as wheat. The virus belongs to the genus Furovirus and is transmitted by the obligate biotrophic root parasite Polymyxa graminis. For the agriculturally important crop barley, no resistances against JSBWMV or its vector P. graminis are known. In this work the infection of the bipartite positive-stranded RNA virus JSBWMV was investigated to understand the interplay between the virus and the host-plant. Nicotiana benthamiana was used as model host. For the viral life cycle the movement of the virus over short distances as well as long distances is important to efficiently establish infection. Viruses encode “movement proteins” (MP) which facilitate the movement processes. For the JSBWMV MP, belonging to the 30K MP-family, a localization to plasmodesmata and to punctuate spots in the plasma membrane was observed. The importance of MPs for intercellular transport of viruses and protection of viral RNA are supported by these results. At both cellular localizations a self-interaction was revealed, which was not shown for a furoviral MP before. To understand the interplay of the virus with the plant cell, interacting RNA and proteins were identified using co-immunoprecipitation (IP) techniques. The experiments showed that the MP interacts with RNAs, which potentially encode transcription factors. Two interacting plant proteins were identified and the interaction was confirmed in further experiments. The potential P-type “pentatricopeptiderepeat-containing protein” (PPR) localized to two distinct subcellular localizations. The C-terminal tagged PPR localized to the chloroplast, while a distribution to the cytoplasm and to plasmodesmata was observed for the N-terminal tagged PPR. RNA-IPs identified interacting RNA-molecules and indicated that the PPR-protein possibly plays a role in regulating mRNA in the chloroplast. The “heat shock protein 70” (HSP70) is a chaperone and experiments displayed a cytoplasmic and nuclear localization. The protein is upregulated in response to the over-expression of the JSBWMV MP and RNA-IP-experiments highlight the role of HSP70 in the plant defense. Interestingly, RNAs interacting with both, MP and HSP70 allowed establishing a link between these two proteins and beta-1,3-glucosidase, one of the main regulators of plasmodesmata size exclusion limit. The results obtained for the JSBWMV MP and the host-plant interacting proteins PPR and HSP70 allow a deeper insight into the interaction of MP with the host cell in the viral infection. This knowledge may in the future be developed into new resistance strategies against JSBWMV.

This dissertation was only published as a printed edition.