Dengue Virus Capsid Protein Localization and Its Interplay with Viral RNA Synthesis

ANDREA GAMARNIK

Janelia Farm, Virginia, USA

Conferencia; HHMI meeting on RNA Chemistry Functions and Subcellular Trafficking; 2009

Howard Hughes Medical Institute

Dengue Virus Capsid Protein Localization and Its Interplay with Viral RNA Synthesis / Andrea Gamarnik, Fundación Instituto Leloir-CONICET, Buenos Aires, Argentina The genus Flavivirus comprises a large group of emerging and re-emerging pathogens capable of causing severe human diseases. It includes dengue (DENV), yellow fever, West Nile, tick borne encephalitis, and Japanese encephalitis viruses. The viral genome is a single plus-stranded RNA molecule that serves as messenger for viral protein synthesis, template for RNA amplification, and substrate for encapsidation. The mechanism by which the utilization of the viral RNA is regulated during infection remains unclear. We have previously identified an RNA element in the viral genome that provides specificity for the viral polymerase. This element, known as SLA, recruits the polymerase and promotes RNA synthesis in a circularized genome. A number of cis-acting elements were found to regulate this process. In contrast, little is known about the recognition of the viral RNA by the capsid (C) protein during genome encapsidation. To study the process of DENV encapsidation, the subcellular localization of the C protein in infected cells was first investigated. Previous reports indicated that the C protein accumulates in the nucleus, however, using different techniques to preserve cellular membranes, the C protein was observed in ring-like patterns in the cytoplasm. A specific accumulation of the C protein surrounding organelles known as lipid droplets (LDs) was observed. LDs are dynamic structures derived from the ER that store neutral lipids and play crucial roles in lipid metabolism. Interestingly, viral infection resulted in an increase in the size and the number of LDs, suggesting a link between LD metabolism and DENV replication. Manipulation of infectious clones and generation of new reporter DENVs allowed us to define the molecular basis of C protein association to LDs. Specific amino acids on the alfa- helix, located in the center of the C protein, were found to be crucial for both accumulation of C on LDs and DENV infectious particle formation. Although we found that the C protein was dispensable for efficient viral RNA amplification, mislocalization in the cytoplasm of a mutated C protein decreased viral RNA synthesis.  We propose that LDs play multiple roles during the viral life cycle; they could sequester the viral C protein early during infection to ensure efficient RNA synthesis and provide a scaffold for genome encapsidation.