Dynamic RNA Structures Involved in Dengue Virus Genome Amplification

ANDREA GAMARNIK

MEXICO

Congreso; Congreso Nacional de Virología, MEXICO, Merida; 2009

Dynamic RNA Structures Involved in Dengue Virus Genome Amplification / Sergio Villordo, Claudia Filomatori, Diego Alvarez, Gabriel Iglesias, and Andrea Gamarnik Fundación Instituto Leloir, Buenos Aires, Argentina Dengue virus is an important human pathogen transmitted by mosquitoes that belongs to the Flaviviridae family. The viral genome is a single stranded RNA molecule of positive polarity that is amplified by a two-step process. First, the RNA dependent RNA polymerase (RdRp), encoded in the viral protein NS5, makes a complementary strand with negative polarity, which then, serves as a template to copy multiple viral genomes. We have previously identified an RNA structure present at the 5’ end of the genome (named SLA) that binds the polymerase and promotes viral RNA synthesis. This SLA promoter functions together with two pairs of complementary sequences located at the ends of the genome that mediate RNA cyclization. In the proposed model, genome cyclization facilitates the initiation of RNA synthesis by locating the RdRp (bound to the 5’SLA) near the 3’ end initiation site. Using in vitro RNA polymerase assays and infectious viral RNAs, the structural elements required for SLA activity and viral replication were recently defined. These elements were found to be conserved among different members of the Flavivirus genus, providing evidence for a common mechanism of promoter mediated RNA synthesis. Moreover, we investigated the role of linear and circular conformation of the viral genome during dengue virus replication. Incorporation of mutations in infectious clones that increase the stability of the circular or the linear forms of the RNA resulted in spontaneous mutations that restored the wild type stability of the long-range RNA-RNA interaction.  We conclude that the dengue virus genome is dynamic and both linear and circular conformations are necessary for RNA amplification.