Rasputin/G3BP mediates o’nyong-nyong virus subversion of antiviral immunity in Anopheles coluzzii

Abstract

ABSTRACTThe G3BP proteins in vertebrates and Aedes mosquito ortholog, Rasputin, are essential for alphavirus infection, but the underlying mechanism of Rasputin/G3BP proviral activity is poorly understood. It has been suggested that G3BP could influence host immune signaling, but this has not been functionally demonstrated. Here, we find that depletion of Rasputin activity in Anopheles mosquitoes, the primary vectors of the alphavirus o’nyong-nyong (ONNV), provokes dysregulation of the antiviral Imd, JAK/STAT and RNAi pathways, indicating that Rasputin is required for expression of normal basal immunity in uninfected mosquitoes. Depletion of Rasputin during ONNV bloodmeal infection causes increased transcript abundance of genes in the Imd pathway including positive regulator Rel2, and decreases ONNV infection in mosquitoes. Loss of Rasputin is complemented by co-depletion of Imd pathway positive regulator, Rel2, which restores normal ONNV infection levels. Thus, the presence of Rasputin is required for ONNV inhibition of Imd activity, and viral inhibition of Imd explains much of the Rasputin proviral activity. The viral non-structural protein 3 (nsP3) binds to Rasputin and alters the profile of cellular proteins binding to Rasputin. In the presence of nsP3, 48 Rasputin-binding proteins are unchanged but seven binding proteins are excluded and eight new proteins bind Rasputin. The Rasputin binding partners altered by nsP3 are candidate factors for ONNV immune manipulation and subversion through Rasputin. Overall, these results are consistent with and strongly suggest a mechanism in which ONNV, probably nsP3, co-opts the normal Rasputin function assuring basal cellular immune activity in order to inhibit antiviral immunity and promote infection. These observations may be generalizable for Rasputin function during alphavirus infection of other mosquitoes, as well as for G3BP function in the mammalian host, and could offer a target for vector-based control of arbovirus transmission.