Two viral proteins translated from one open reading frame target different layers of plant defense

Multilayered defense responses are activated upon pathogen attack. Viruses utilize a number of strategies to maximize the coding capacity of their small genomes and produce viral proteins for infection, including suppression of host defense. Here, we reveal translation leakage as one of these strategies: two viral effectors encoded by tomato golden mosaic virus, chloroplast-localized C4 (cC4) and membrane-associated C4 (mC4), are translated from two in-frame start codons and function cooperatively to suppress defense. cC4 localizes in chloroplasts, to which it recruits NbPUB4 to induce ubiquitination of the outer membrane; as a result, this organelle is degraded, and chloroplast-mediated defenses are abrogated. However, chloroplast-localized cC4 induces the production of singlet oxygen (¹O₂), which in turn promotes translocation of the ¹O₂ sensor NbMBS1 from the cytosol to the nucleus, where it activates expression of the CERK1 gene. Importantly, an antiviral effect exerted by CERK1 is countered by mC4, localized at the plasma membrane. mC4, like cC4, recruits NbPUB4 and promotes the ubiquitination and subsequent degradation of CERK1, suppressing membrane-based, receptor-like kinase-dependent defenses. Importantly, this translation leakage strategy seems to be conserved in multiple viral species and is related to host range. This finding suggests that stacking of different cellular antiviral responses could be an effective way to abrogate viral infection and engineer sustainable resistance to major crop viral diseases in the field.