Dengue Virus Capsid Functions: Encapsidation and Uncoating

BYK, L; IGLESIAS, N.G.; SAMSA, M.M.; GEBHARD, L.; DE BORBA. L.; VILLORDO, S.M.; GAMARNIK, A.V.

Simposio; Keystone Symposium The Ins and Outs of Viral Infection: Entry, Assembly, Exit and Spread; 2014

Dengue virus is an important human pathogen transmitted by mosquitoes. It has been recently estimated a global burden of 390 million infections per year, while no licensed vaccines or specific antivirals exist. Dengue and other flavivirus capsid proteins form a nucleocapsid with the viral RNA genome. This ribonucleoprotein complex is incorporated into the viral particle during encapsidation and is disassembled during uncoating. Although RNA encapsidation and uncoating are essential processes for viral infection, the mechanisms by which the capsid protein recruits and frees the viral genome are largely unknown. We examined determinants in the dengue virus capsid protein necessary for encapsidation and uncoating. The capsid coding sequence contains a number of RNA structures that modulate viral RNA replication, which hampers detailed studies on amino acids involved in encapsidation. These elements were confirmed by chemical probing using SHAPE analysis and their functions defined by specific deletions. Based on this information, reporter viruses were designed in which the RNA replication signals were separated from the capsid coding region. A systematic mutational analysis using this system indicated that a high density of basic residues at the N-terminus and at the center of the alpha-4 helix, rather than specific amino acids in defined positions, were required for infectious particle production. In contrast, specific amino acids were found to reduce slightly the amount of infectious particles but completely abrogated viral infectivity. These viruses were able to enter the susceptible cell, but translation and replication were impaired. New ideas and requirements for dengue virus encapsidation and uncoating will be discussed.