Please use this identifier to cite or link to this item: http://scholarbank.nus.edu.sg/handle/10635/19073
Title: Studies on interactions between HCRSV coat protein and host proteins from kenaf
Authors: ZHANG XIN
Keywords: Sulfite Oxidase, HCRSV CP, Interaction, cDNA library
Issue Date: 20-Jul-2010
Source: ZHANG XIN (2010-07-20). Studies on interactions between HCRSV coat protein and host proteins from kenaf. ScholarBank@NUS Repository.
Abstract: Hibiscus chlorotic ringspot virus (HCRSV) infection can cause severe chlorotic ringspot symptoms and stunted growth on Hibiscus which is an ornamental plant grown in many parts of the world. HCRSV coat protein (CP) is an important viral gene product required for encapsidation and viral systemic movement. To better understand the roles of HCRSV CP in viral infection and its interactions with host proteins, a kenaf cDNA library was constructed and screened using a yeast two-hybrid (Y2H) system to identify CP-interacting proteins. Using the Y2H system, several putative proteins that interact with HCRSV coat protein were identified. These proteins include sulfite oxidase (SO), a putative major latex-like protein, a putative chaperon P13.9, a C2 domain-containing protein, a ricin domain-containing protein and putative alpha-D-xylosidase like protein. The interaction of CP and SO was confirmed by in vitro binding assay. Bimolecular fluorescence complementation (BiFC) assay was used to colocalize the two proteins in vivo. The interaction was found to be associated with peroxisomes by immunofluorescent labeling using anti serine-lysine-leucine (SKL) signal peptide antibody. A second Y2Hstudy showed that both the P and S domains of CP could interact with SO. This is probably due to the exposure of these two domains on the outer surface of the capsid. In addition, Transmission electron microscopy (TEM) revealed that peroxisomes were aggregated in HCRSV infected cells and the CP was localized within the peroxisomes by immuno-gold labeling. Biochemical assays of the total protein from kenaf leaf extracts showed that SO activity and SO-dependent H2O2-generating activity in the HCRSV-infected leaves increased compared to the mock-inoculated kenaf plants. Thus, it is speculated that the interaction of HCRSV CP and SO is important for the plant to up-regulate the sulfate level and SO activity, which may be responsible for the virus accumulation or plant defense during the virus infection. Taken together, it is shown that HCRSV infection upregulates plant sulfite oxidase (PSO) transcripts and increases sulfate levels in kenaf. A model is proposed on how HCRSV is able to overcome plant resistance responses to establish infection in plants. The interaction between ribosomal protein L10A and HCRSV CP was also further studied (Chapter 5).
URI: http://scholarbank.nus.edu.sg/handle/10635/19073
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