Characterization of RNA aptamers directed against the nucleocapsid protein of Rift Valley fever virus

Nucleocapsid protein (N) is an essential RNA binding protein in many RNA viruses. During replication, N protein encapsidates viral genomic and antigenomic RNA, but not viral mRNA or other cellular RNAs. To discriminate between different species of RNA in a host cell, it is likely that N interacts with specific sequences and/or secondary structures on its target RNA. In this study, we explore the RNA binding properties of N using both natural and artificially selected RNAs as ligands. We found that N binds to RNAs that resemble the terminal panhandle structures of RVFV genomic and antigenomic RNA. Furthermore, we used SELEX to isolate RNA aptamers that bound N with high affinity and determined that N specifically recognizes and binds to GAUU and pyrimidine/guanine motifs. Interestingly, BLAST analysis revealed the presence of these motifs within the coding region of the viral genome, suggesting that N may interact with non-terminal viral RNA sequences during replication. Finally, the aptamer RNAs were used to construct a sensitive fluorescence based sensor of N binding with potential applications for drug screening and imaging methodologies.

► In this study, we use SELEX to derive high affinity RNA ligands to Rift Valley fever virus nucleocapsid protein. ► The sequences of the RNA aptamers inform about the RNA sequence and structural preferences of nucleocapsid protein. ► We constructed a fluorescent biosensor that reports on nucleocapsid-RNA interactions. ► This biosensor can be used as a molecular target assay for potential new antiviral drug screening.