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

Tomato bushy stunt virus (TBSV) encodes a small gene (p19) nested within th e cell-to-cell movement gene (p22), and their translation yields two protei ns with separate activities for virus spread and symptom induction. The obj ective of this study was to determine the biological relevance associated w ith the translational mechanism responsible for expression of the nested p2 2 and p19 genes. Introduction of site-specific mutations to optimize the tr anslational start site context of p22 caused a substantial shift in the rat io of the two proteins, mainly because it dramatically reduced the otherwis e 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 affec t virus replication or movement in Nicotiana spp, that support a systemic i nfection. In contrast, bio-assays with hypersensitive Nicotiana hosts illus trated that a substantially elevated p22/p19 protein ratio increased the si ze of p19 protein-mediated lesions whereas those induced by the p22 protein tended to be smaller. The reduced levels of p19 protein prevented the onse t of a lethal apical necrosis in systemically infected Nicotiana benthamian a plants. Furthermore, the increased p22/p19 protein ratio impaired the abi lity of TBSV to systemically invade spinach plants. These results suggest t hat control of tombusvirus p22 and p19 protein ratios and dosage through co ntext-dependent leaky scanning provides a co-translational mechanism to coo rdinate their biological activities.