Scott Tersteeg, University of Lethbridge

Scott Tersteeg, Anneke De Klerk, and Trushar Patel (University of Lethbridge, Lethbridge, Canada)

Expressing & Multiplying

Flaviviridae are a family of single-stranded, positive-sense, RNA viruses of global concern due to recent outbreaks of Zika and Dengue infection causing debilitating and fatal effects in people across numerous equatorial and tropical global south nations. As a member of the Flaviviridae family, Japanese Encephalitis Virus (JEV) has highly structured and functional 5' and 3' Terminal Regions (TRs) which aid in viral genome cyclization, host immune evasion, viral RNA synthesis, and transcription through the recruitment of the non-structural protein 5 (NS5) which is responsible for negative and positive strand synthesis. A promoter for the NS5, known as Stem-Loop A (SLA), has been determined to be located on the 5' TR. Despite negative-strand synthesis being initiated on the 3' TR, the mechanism of translocation of NS5 from one TR to the other is unknown but is assumed to be intricately linked. There are currently several models proposed by which NS5 begins negative-stand synthesis that we aim to assess. Using small angle X-ray scattering (SAXS) we have obtained a solution structural model for NS5 as a dimer as well as binding data of NS5 to each TR independently. In conjunction with kinetic data determining the rate of association and dissociation of NS5-TR complexes under proposed model conditions, we seek to apply this data to gain a deeper understanding of how negative-strand Flaviviridae synthesis begins.