Toscana virus non-structural protein NSs acts as E3 ubiquitin ligase promoting RIG-I degradation

It is known that the non-structural protein (NSs) of Toscana virus (TOSV), an emergent sandfly-borne virus causing meningitis or more severe central nervous system injuries in humans, exerts its function triggering RIG-I for degradation in a proteasome-dependent manner, thus breaking off the IFN-β production. The non-structural protein of different members of Bunyavirales has recently appeared as a fundamental protagonist in immunity evasion through ubiquitination-mediated protein degradation targets. We showed that TOSV NSs has an E3 ubiquitin ligase activity, mapping at the carboxy-terminal domain and also involving the amino-terminal of the protein. Indeed, neither the amino- (NSsΔN) nor the carboxy- (NSsΔC) terminal-deleted mutants of TOSV NSs were able to cause ubiquitin-mediated proteasome degradation of RIG-I. Moreover, the addition of the C-terminus of TOSV NSs to the homologous protein of the Sandfly Fever Naples Virus, belonging to the same genus and unable to inhibit IFN-β activity, conferred new properties to this protein, favoring RIG-I ubiquitination and its degradation. NSs lost its antagonistic activity to IFN when one of the terminal residues was missing. Therefore, we showed that NSs could behave as an atypical RING between RING (RBR) E3 ubiquitin ligases. This is the first report which identified the E3 ubiquitin ligase activity in a viral protein among negative strand RNA viruses.