Analysis of promoter binding proteins of Ebp1 that is inhibitor protein of influenza virus RNA polymerase

Abstract

Ebp1 is a member of the PA2G4 family protein and initially isolated as an ErB3 (an epidermal receptor tyrosine kinase) binding protein. Ebp1 inhibits cell growth and repress transcription of E2F-regulated cell cycle genes. Previously, we reported that Ebp1 protein interacted with RNA polymerase subunit PB1 of the influenza virus and disturbed in vitro RNA synthesis by influenza RNA polymerase. Recently we found that after influenza virus infection to the cell, Ebp1 expression is induced at 4 hours after viral infection. By using reverse genetics method, Ebp1 inhibits influenza virus replication. Ebp1 expression mechanism is very interesting because the expression of Ebp1 is earlier than that of virus proteins. In addition, in uninfection cell, Ebp1 is expressed from G1 to S phase in cell cycle-dependent manner. Therefore we study to identify the transcription factor of Ebp1 and understand the Ebp1 expression mechanism by influenza viral infection. Ebp1 promoter region was cloned into pTurboGFP. A pTurboGFP-Ebp1 promoter plasmid was mixed with nucleus extract form infection / uninfection cell. The binding proteins were eluted and analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). We identified several candidate proteins.