Mapping of the RNA-binding domain of the cucumber mosaic virus movement protein (original) (raw)
Related papers
Archives of Virology, 1996
The complete nucleotide sequence of RNA 3 of a Spanish isolate of cucumber mosaic virus (CMV-24) has been determined. The encoded putative cell-to-ceil movement protein (3a protein) and the coat protein are 279 and 218 amino acids long, respectively. The 3a protein was expressed in Escherichia coti using the vector pT7-7 and was used to raise an immunoserum. We have followed the time course of accumulation of the 3a protein, in parallel to that of the coat protein, and its subcellular localization as a function of time after CMV-24 infection on tobacco plants. The maximum accumulation level of the 3a protein was reached at early stages of infection, being detected in the cytosolic and the cell wall fractions. At later stages of infection, a decline in accumulation levels of the 3a protein was observed, and the protein was essentially associated with the cell wall fractions. These data were corroborated by immunocytochemistry performed in both infected and 3a-expressing transgenic tobacco plants.
Phosphorylation of the movement protein of Cucumber mosaic virus in transgenic tobacco plants
2002
The 3a protein of Cucumber mosaic virus is essential for the cell-to-cell movement of the viral RNA through plasmodesmata. We have introduced an epitope peptide before the stop codon of the 3a protein and cloned the tagged ORF into a binary vector for Agrobacterium-mediated transformation. The established transgenic tobacco lines produced the 3a protein, which was specifically detected with anti-3a and anti-epitope antisera. Metabolic labeling and subsequent immunoprecipitation revealed that [ 32 P]-orthophosphate was incorporated into the 3a protein. The phosphoamino acid analysis indicated that the 3a protein contained phosphoserine but not phosphothreonine or phosphotyrosine. This is the first demonstration of the 3a protein phosphorylation in planta.
Journal of General Virology
The capsid protein (CP) of Cucumber mosaic virus (CMV) is required for cell-to-cell movement, mediated by the 3a movement protein (MP). Deletion of the C-terminal 33 amino acids of the CMV 3a MP (in the mutant designated 3aDC33 MP) resulted in CP-independent cell-to-cell movement, but not long-distance movement. RNA-binding studies done in vitro using isolated bacterially expressed MP showed that the 3aDC33 MP bound RNA more strongly, with fewer regions sensitive to RNase and formed cooperatively bound complexes at lower ratios of protein : RNA than the wild-type (wt) 3a MP. Analysis of the architecture of the complexes by atomic force microscopy showed that the wt 3a MP formed a single type of complex with RNA, resembling beads on a string. By contrast, the 3aDC33 MP formed several types of complexes, including complexes with virtually no MP bound or thicker layers of MP bound to the RNA. Assays showed that protein-RNA complexes containing high levels of either MP inhibited the infectivity and in vitro translatability of viral RNAs. The 3aDC33 MP inhibited these processes at lower ratios of protein : RNA than the wt 3a MP, consistent with its stronger binding properties. The apparent contradiction between these inhibition data and the CP-independent cell-to-cell movement of CMV expressing the 3aDC33 MP is discussed.
Journal of General Virology, 2004
The capsid protein (CP) of Cucumber mosaic virus (CMV) is required for cell-to-cell movement, mediated by the 3a movement protein (MP). Deletion of the C-terminal 33 amino acids of the CMV 3a MP (in the mutant designated 3aDC33 MP) resulted in CP-independent cell-to-cell movement, but not long-distance movement. RNA-binding studies done in vitro using isolated bacterially expressed MP showed that the 3aDC33 MP bound RNA more strongly, with fewer regions sensitive to RNase and formed cooperatively bound complexes at lower ratios of protein : RNA than the wild-type (wt) 3a MP. Analysis of the architecture of the complexes by atomic force microscopy showed that the wt 3a MP formed a single type of complex with RNA, resembling beads on a string. By contrast, the 3aDC33 MP formed several types of complexes, including complexes with virtually no MP bound or thicker layers of MP bound to the RNA. Assays showed that protein-RNA complexes containing high levels of either MP inhibited the infectivity and in vitro translatability of viral RNAs. The 3aDC33 MP inhibited these processes at lower ratios of protein : RNA than the wt 3a MP, consistent with its stronger binding properties. The apparent contradiction between these inhibition data and the CP-independent cell-to-cell movement of CMV expressing the 3aDC33 MP is discussed.
Heterologous Movement Protein Strongly Modifies the Infection Phenotype of Cucumber Mosaic Virus
Journal of Virology, 2002
A hybrid virus (CMVcymMP) constructed by replacing the movement protein (MP) of cucumber mosaic cucumovirus (CMV) with that of cymbidium ringspot tombusvirus (CymRSV) was viable and could efficiently spread both cell to cell and long distance in host plants. The hybrid virus was able to move cell to cell in the absence of functional CP, whereas CP-deficient CMV was restricted to single inoculated cells. In several Chenopodium and Nicotiana species, the symptom phenotype of the hybrid virus infection was clearly determined by the foreign MP gene. In Nicotiana debneyi and Nicotiana tabacum cv. Xanthi, the hybrid virus could move systemically, contrary to CymRSV.
Molecular Plant-Microbe Interactions®, 1997
Systemic spread of tobacco mosaic virus (TMV) that lacks a functional movement protein (TMVΔMP) was investigated in grafted tobacco (Nicotiana tabacum) plants. Transgenic plants that express the 30-kDa movement protein (MP) gene (MP) under the control of the rolC (phloem-specific) or pal2 (xylem-specific) promoters were unable to support systemic infection by the mutant virus, while plants that express the MP gene from the cauliflower mosaic virus 35S promoter (35S:MP) led to systemic infection. Doubly grafted plants were constructed in which plants containing the 35S:MP gene were used as root stock and plants carrying various MP constructs constituted the middle scion. The upper scion contained the 35S:MP gene in plants that produce a hypersensitive response when systemically infected by TMV. TMVΔMP moved systemically and produced complete necrosis in the upper scion when expression of MP in the middle scion was under the control of the rolC or 35S promoter, but not when the pal2 p...
Cell-to-cell movement of three genera (+) ss RNA plant viruses
Acta Physiologiae Plantarum, 2011
The current investigations of three genera plant virus cell-to-cell movement were presented. Viruses reveal different local transport strategies, but all of them are the results of virus factors-host components interactions. The Tobacco mosaic virus (TMV) does not require capsid protein for translocation through plasmodesmata but 30 K movement protein participates in this process. It was found direct or indirect TMV movement proteins host partners in Tobamovirus movement like: pectin methylesterase, movement protein binding 2C, chaperones or cytoskeleton components and endoplasmatic reticulum membranes. The Potex-and Potyvirus cell-to-cell movement is closely related to replication network. The PVX capsid protein and triple gene block protein system are responsible for efficient local transport. Potyviruses move through the plasmodesmata by involving viral encoded proteins but not specific movement proteins. While the Potyvirus is the biggest known plant virus genus, host components participating in or regulating directly its plasmodesmata-movement are still not clear.