Identification of Receptor Transporting Proteins As Conserved Antiviral Effectors in Vertebrates

Viruses and their hosts are engaged in "genetic arms races" in which each side attempts to gain the advantage over evolutionary time. Results of these conflicts are wide-ranging: viruses diversify, hosts establish species-specific barriers to some viruses while remaining susceptible to others, and the lines for future genetic conflicts are drawn. In mammals, many antiviral effectors -- proteins that directly inhibit viral infection -- show species- or lineage-specific properties which are believed to be the result of past or ongoing conflicts. Bats harbor a greater diversity of viruses than any other mammalian order, and a growing body of research has described unique adaptations in bats that are in part responsible for, and perhaps a response to, this unique status. We hypothesized that the frequent encounters between bats and viruses would drive unique adaptations in the antiviral effectors that serve on the front lines of virus-host genetic conflicts. We identified RTP4 from the bat Pteropus alecto as a potent inhibitor of flavivirus infection. Mechanistic studies determined that RTP4 is an RNA-binding protein that associates with flavivirus replication machinery, binds replicating viral RNA, and suppresses viral genome amplification. Phylogenomic analysis revealed that RTP4 has evolved under positive selection in several mammalian lineages, consistent with a model in which host-virus conflicts have shaped its evolution as a restriction factor not only in bats but across mammals. We assessed the antiviral efficacy of diverse mammalian RTP4 orthologs and found that orthologs exhibit striking patterns of antiviral specificity. Further highlighting the specificity of the host-virus arms race, experimental evolution demonstrated that a flavivirus can mutate to escape RTP4-imposed restriction in a species-specific manner. In follow-up work, we identified signatures of positive selection in several non-mammalian RTP homologs, indicative of a putative role in innate immunity. We screened a collection of vertebrate RTPs against a panel of viruses and identified antiviral RTPs in the African clawed frog, Xenopus laevis. These antiviral Xenopus RTPs exhibit mosaic phenotypes that resemble those of mammalian RTP4 orthologs. Within the context of our findings with mammalian RTP4, these data suggest that Receptor Transporter Proteins are involved in host-virus genetic conflicts outside of Mammalia.