EPS8 facilitates uncoating of influenza A virus

Gloria P. Larson; Vy Tran; Shuiqìng Yú; Yíngyún Caì; Christina A. Higgins; Danielle M. Smith; Steven F. Baker; Sheli R. Radoshitzky; Jens H. Kuhn; Andrew Mehle

The structure of the NCI-60 screen and our data indicated EPS8 functions in early stages of FLUAV replication. We therefore conducted a series of experiments to determine where in the viral replication cycle EPS8 functioned to enhance infection ( Figure 3A ). We first assessed whether EPS8 affects infection through a mechanism that directly impacts viral polymerase activity. Polymerase activity was reconstituted in the absence of infection by expressing the heterotrimeric viral polymerase subunits PA, PB1, and PB2, nucleoprotein (NP), and a vRNA-like reporter encoding firefly luciferase. Polymerase activity was not statistically different in the presence or absence of exogenous EPS8 ( Figure 3B ). Immunoblotting confirmed high levels of exogenously expressed EPS8. This finding establishes that EPS8mediated enhancement of viral gene expression is not due to direct impacts on the viral polymerase but rather an upstream step in the early stages of infection.