Two mutually exclusive evolutionary scenarios for allexiviruses that overcome host RNA silencing and autophagy by regulating viral CRP expression

The genus Allexivirus currently includes eight virus species that infect allium plants. Previously, we showed that there are two distinct groups of allexiviruses (deletion [D]-type and insertion [I]-type) based on the presence or absence of a 10- to 20-base insert (IS) between the coat protein (CP) and cysteine rich protein (CRP) genes. In the present study of CRPs to analyze their functions, we postulated that evolution of allexiviruses may have been largely directed by CRPs and thus proposed two evolutionary scenarios for allexiviruses based mainly on the presence or absence of IS and determined by how the allexiviruses challenge host resistance mechanisms (RNA silencing and autophagy). We found that both CP and CRP are RNA silencing suppressors (RSS), that they can inhibit each other's RSS activity in the cytoplasm, and that CRP becomes a target of host autophagy in the cytoplasm but not CP. To mitigate CRP interference with CP, and to increase the CP's RSS activity, allexiviruses developed two strategies: confinement of D-type CRP in the nucleus and degradation of I-type CRP by autophagy in the cytoplasm. Here, we demonstrate that viruses of the same genus achieve two completely different evolutionary scenarios by controlling expression and subcellular localization of CRP. Author summaryAllexiviruses in the genus Allexivirus mainly infect allium plants and are often found as mixed infections in garlic. The genus currently includes 7 allexiviruses, with diverse sequence differences even among isolates within the same species. In plant viruses, RNA silencing suppressors (RSSs) play an important role in viral host adaptation by acting as virulence determinants, and have been found to be the driving force of viral evolution. The allexivirus protein, CRP has been found to be an RSS. The phylogenetic analysis based on this gene well supported the classification of the 7 allexiviruses, and the same results were obtained using the coat protein (CP) gene. In addition, allexiviruses are divided into two groups depending on the presence or absence of an inserted sequence of 10 to 20 nucleotides between the CP and CRP genes. Because a balance between CP and CRP levels seemed to be important for viral pathogenicity, we investigated whether CP has some effect on the RSS activity of CRP, and found that CP was also an RSS. Surprisingly, we found that CP and CRP inhibit each other's RSS activity; this is the first report on an antagonistic interaction between two RSSs. We here explain that allexiviruses have chosen two totally different evolutionary scenarios for their survival depending on how they maintain their RSS activity in infected cells.