[Identification of the Transcriptional Activity Domain of EV71 3D(pol)].

Abstract

Enterovirus 71（EV71）is one of the major pathogens of hand, foot and mouth disease (HFMD). The EV71 genome encodes an RNA-dependent RNA polymerase（RdRp）,3D(pol),which is critical for genome transcription and translation. However, how the 3D(pol) interacts with the host remains unclear. Yeast two-hybrid systems provide an effective approach for detecting protein-protein interactions. In this report, we inserted the DNA sequence of 3D(pol) into the pGBKT7 vector as the bait plasmid for the yeast two-hybrid experiment and transformed the plasmid into the yeast AH109 strain. We detected the expression,cytotoxicity and self-activity of 3D(pol).The 3D(pol) expressed well without affecting cell growth but exhibited strong transcriptional activation in yeast cells. We further constructed a series of pGBKT7-3D(pol) deletion mutants and identified the shortest transcriptional activation domain（1-94aa）using a self-activation assay. The results provide a molecular basis for screening the host proteins that interact with 3D(pol) using the yeast two hybrid system.