Biological activity of two tombusvirus proteins translated from nested genes is influenced by dosage control via context-dependent leaky scanning

Abstract

Tomato bushy stunt virus (TBSV) encodes a small gene (p19) nested within the cell-to-cell movement gene (p22), and their translation yields two proteins with separate activities for virus spread and symptom induction. The objective of this study was to determine the biological relevance associated with the translational mechanism responsible for expression of the nested p22 and p19 genes. Introduction of site-specific mutations to optimize the translational start site context of p22 caused a substantial shift in the ratio of the two proteins, mainly because it dramatically reduced the otherwise abundant levels of p19 protein accumulation in vitro and in vivo. Changes in the dosage or ratios of p22 and p19 proteins failed to noticeably affect virus replication or movement in Nicotiana spp. that support a systemic infection. In contrast, bio-assays with hypersensitive Nicotiana hosts illustrated that a substantially elevated p22/p19 protein ratio increased the size of p19 protein-mediated lesions whereas those induced by the p22 protein tended to be smaller. The reduced levels of p19 protein prevented the onset of a lethal apical necrosis in systematically infected Nicotiana benthamiana plants. Furthermore, the increased p22/p19 protein ratio impaired the ability of TBSV to systematically invade spinach plants. These results suggest that control of tombusvirus p22 and p19 protein ratios and dosage through context-dependent leaky scanning provides a co-translational mechanism to coordinate their biological activities.