Abstract 1782 Identifying Essential RNA Structures for Noncanonical Cap-Independent Translation in Blackcurrant Reversion Virus

RNA structures located in the untranslated regions (UTRs) of viral RNA genomes are required for viral replication and pathogenesis, including noncanonical translation initiation.The 3 0 UTR of plant RNA viruses, such as the blackcurrant reversion virus (BRV), contain a variety of structures that function as cap-independent translation enhancers (CITEs).These CITEs hijack a subset of translation factors to initiate viral protein synthesis in a cellular environment where capdependent translation initiation is repressed.Both genomic RNAs in BRV contain a CITE, yet the putative stem-loop 4 (SL4) is not conserved between them, raising the question; is SL4 essential for BRV translation initiation?Minimum freeenergy structural modeling revealed that deleting SL4 has negligible impact on the overall CITE secondary structure.Initial in vitro transcription and reporter translation assays show a modest effect on translation initiation when SL4 is deleted, suggesting SL4 is not critical for BRV RNA-1 function.Our results were further supported by multiple sequence alignment of viral variants showing minimal conservation in the loop of SL4.Collectively, our data suggests that SL4 structure folds independently of the other CITE stem-loops, and it may play a minor or redundant role in noncanonical translation initiation.