Browsing by Subject "rabE1"

The leaves of healthy potato (Solanum tuberosum L.) plants and leaves systemically infected with Potato virus A (PVA; a monopartite positive sense single stranded RNA genome, genus: Potyvirus, family: Potyviridae) have been previously analysed and compared for the nuclear proteome to better understand the role of nucleus in plant virus infection. The analysis indicated that a small GTP-binding protein (ras-related protein; rabE1) is found in the nuclei of PVA-infected leaves but is absent from nuclei of healthy leaves. Therefore, to confirm the differential localization, this study was conducted. Subcellular localization of proteins was studied by tagging the protein with a marker protein, green fluorescent protein (GFP). The mRNA of ras-like protein, rabE1, of potato cv. Pentland Crown was cloned and expressed with GFP fused to the N- or C-terminus of rabE1 (GFP-ras and ras-GFP respectively). Plants of Nicotiana benthamiana were inoculated with PVA by two methods. Firstly, the Agrobacterium strains expressing GFP-ras or ras-GFP were mixed with the Agrobacterium strain expressing red fluorescent protein (RFP) or PVA-RFP and introduced to the healthy leaves of N. benthamiana plants and observed under epifluorescence microscope three days post inoculation. Secondly, PVA-RFP was agroinoculated to two or three basal leaves of young N. benthamiana plants and plants were grown for 12 days to allow systemic infection. Thereafter, PVA infected upper leaves and healthy leaves were infiltrated with GFP-ras or ras-GFP and the leaves were observed under epifluorescence microscope three days after infiltration. In healthy plants, GFP-ras localized to the cytoplasm and ras-GFP localized mostly to the cytoplasm and in few cells it was found both in the nucleus and cytoplasm. Whereas, in PVA infected cells, GFP-ras localized to the cytoplasm and in some cells to both nucleus and cytoplasm suggesting that PVA infection increases GFP-ras localization to the nucleus. However, no difference was observed in localization of ras-GFP in PVA infected and healthy leaves. The hypothesis of the research is thus partially true. Our study shows that rabE1 can be differentially localized in the healthy and PVA-infected plant cells.