Critical elements of dengue virus translation

Dengue virus (DEN) is the most prevalent arthropod-borne virus and a major public health threat worldwide. Despite the worldwide morbidity and mortality associated with DEN infection, neither the molecular virology nor the pathogenesis is well understood. Therefore, the focus of these studies, initially, was to characterize critical elements of the DEN lifecycle.; Efforts to determine DEN cellular tropism in vitro revealed a disparity in the ability of closely related DEN strains from Thailand and Nicaragua to productively infect primary human dermal fibroblasts and myeloid cells. Further dissection of the viral lifecycle indicated that all strains were equally capable of binding, entry, and nucleocapsid penetration, and displayed comparable stability of positive strand viral RNA over time in primary cells. However, the low-passage Nicaraguan isolates were much less efficient in their ability to translate viral proteins. Genetic differences were identified between the 3' untranslated regions (3' UTR) of the Nicaraguan and Thai strains, which, when placed in RNA reporter constructs, directly affected the efficiency of viral translation. These results indicated that the efficiency of translation of input viral RNA was crucial to viral infectivity.; The mechanism of DEN translation has been relatively unexplored. DEN contains a 5' m7GpppN-capped positive-stranded RNA genome with a long, non-polyadenylated (polyA) 3' untranslated region (UTR) that has been presumed to undergo translation in a cap-dependent manner. During cap-dependent translation initiation, the eukaryotic initiation factor eIF4E, a component of the cap-binding complex, is utilized to recognize the cap structure at the 5' end of viral and cellular mRNAs. However, under conditions that inhibit cap-dependent translation by targeting eIF4E, we found that DEN replication and translation were not affected. Additional studies demonstrated a novel mechanism by which the capped DEN genome can also be translated cap-independently under conditions of reduced eIF4E through the use of an IRES whose activity requires the presence of the DEN 5' and 3'UTRs. Ultimately, the report of a capped viral RNA that can alternate between mechanisms of translation in response to cellular environment has far-reaching implications for cellular tropism, viral transmission, vector and host competence, and antiviral strategies.