Abstract
Virus life cycle heavily depends on their ability to command the host machinery in order to translate their genomes. Animal viruses have been shown to interfere with host translation machinery by expressing viral proteins that either maintain or inhibit eIF2α function by phosphorylation. However, this interference mechanism has not been described for any plant virus yet. Prunnus necrotic ringspot virus (PNRSV) is a serious pathogen of cultivated stone fruit trees. The movement protein (MP) of PNRSV is necessary for the cell-to-cell movement of the virus. By using a yeast-based approach we have found that over-expression of the PNRSV MP caused a severe growth defect in yeast cells. cDNA microarrays analysis carried out to characterise at the molecular level the growth interference phenotype reported the induction of genes related to amino acid deprivation suggesting that expression of MP activates the GCN pathway in yeast cells. Accordingly, PNRSV MP triggered activation of the Gcn2p kinase, as judged by increased eIF2α phosphorylation. Activation of Gcn2p by MP expression required a functional Tor1p kinase, since rapamycin treatment alleviated the yeast cell growth defect and blocked eIF2α phosphorylation triggered by MP expression. Overall, these findings uncover a previously uncharacterised function for PNRSV MP viral protein, and point out at Tor1p and Gcn2p kinases as candidate susceptibility factors for plant viral infections.
Highlights
Saccharomyces cerevisiae cells transiently inhibit initiation of protein synthesis under environmental stress to avoid misfolding of proteins which could compromise cell viability
To further analyze the specific MPpnrsv-induced growth interference phenotype, we explored the effect on yeast cell growth of other movement protein (MP) from the 30 K superfamily
Measurement of the yeast cell growth at the exponential phase with or without doxycycline showed that Brome mosaic virus (BMV), Cucumber mosaic virus (CMV) and Tobacco mosaic virus (TMV) MPs expression elicited a growth interference phenotype that was totally absent in the case of Grapevine fanleaf virus (GFLV) MP (Fig. 1E)
Summary
Saccharomyces cerevisiae cells transiently inhibit initiation of protein synthesis under environmental stress to avoid misfolding of proteins which could compromise cell viability. The general control non- repressible 2 protein (Gcn2p) regulates the selective translation of the Gcn4p transcription factor upon nutrient deprivation conditions. This regulation is exerted by four uORFs located in the 59 untranslated region of the GCN4 mRNA, which makes it hypersensitive to eIF2 levels. When there is no amino acid limitation, the levels of active eIF2 are high and the uORFs block translation of GCN4 mRNA. Phosphorylation of eIF2a diminishes the levels of active eIF2 and alleviates inhibition by uORFs, favouring GCN4 mRNA translation as well as blocking general translation [1]. High levels of Gcn4p activate the expression of genes encoding amino acid biosynthesis pathways. Arabidopsis and rice genome analysis revealed that plants apparently encode a single eIF2a kinase orthologue of yeast GCN2 [4,5]
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