Structural studies of the RNA promoter and PB1-F2 protein of the influenza A virus

Abstract

Influenza A virus promoter is recognized by the influenza A virus RNA-dependent RNA polymerase and directs both transcription and replication of the viral RNA genome. Within the sequence of this promoter, flu strains exhibit a natural unique variation, either a U or a C, at the $4^th$ position from the 3`` end. Promoters that contain a C residue (C4 promoter), which are invariably found in genome segments that encode the three RNA polymerase subunits (PB1, PB2, and PA), down-regulate transcription but activate genome replication. Here we have determined the structure of the C4 promoter by NMR spectroscopy and compared it with the structure of the U4 promoter, which was determined previously. The structure of the internal loop in the C4 promoter is similar to that of the U4 promoter. However, the terminal stem of the C4 promoter is strikingly different from that of the U4 promoter. These suggest that the internal loop is important for polymerase binding to the promoter and the terminal stem is crucial for differential regulation of transcription and replication. An abundant immunogenic peptide encoded by the +1 reading frame of the PB1 polymerase subunit was found by Chen et al. in the course of characterizing viral determinants recognized by mouse CD8+ T cells. This peptide derives from a novel conserved 87-residue protein, PB1-F2, which has several unusual features compared with other influenza gene products in addition to its mode of translation. These include; much more variable level of expression in individual infected cells than other viral proteins; rapid degradation by proteasomes and possibly other proteases; and location in mitochondria. Exposure of cells to synthetic version of PB1-F2 induces apoptosis, and also influenza viruses with targeted mutations that interfere with PB1-F2 expression induce less extensive apoptosis in human monocytic cells than those with intact PB1-F2. This protein has the Arg-rich domain of positive charge and the C-termina...