Overlapping Local and Long-Range RNA-RNA Interactions Modulate Dengue Virus Genome Cyclization and Replication

The dengue virus genome is a dynamic molecule that adopts different conformations in the infected cell. Here, using RNA fold-ing predictions, chemical probing analysis, RNA binding assays, and functional studies, we identified new cis-acting elements present in the capsid coding sequence that facilitate cyclization of the viral RNA by hybridization with a sequence involved in a local dumbbell structure at the viral 3 = untranslated region (UTR). The identified interaction differentially enhances viral repli-cation in mosquito andmammalian cells. Dengue virus (DENV) is a member of the Flaviviridae familythat includes other important pathogens such as yellow fever virus (YFV), West Nile virus (WNV), Saint Louis encephalitis vi-rus (SLEV), and Japanese encephalitis virus (JEV). The DENV genome is a plus-stranded RNA molecule that contains a single open reading frame flanked by highly structured 5 = and 3 = un-translated regions (UTRs) (1–3). RNA elements located within these regions are responsible for translation initiation and genome replication (4–7). The 5 = UTR is about 100 nucleotides (nt) long and includes three different elements: (i) stem-loop A (SLA), which is the promoter for viral polymerase binding and activatio