A Complicated Message: Identification of a Novel PB1-Related Protein Translated from Influenza A Virus Segment 2 mRNA

Abstract

Influenza A virus segment 2 is known to encode two polypeptides in overlapping open reading frames: PB1, the polymerase, and PB1-F2, a proapoptotic virulence factor. We show that a third major polypeptide is synthesized from PB1 mRNA via differential AUG codon usage. PB1 codon 40 directs translation of an N-terminally truncated version of the polypeptide (N40) that lacks transcriptase function but nevertheless interacts with PB2 and the polymerase complex in the cellular environment. Importantly, the expression of N40, PB1-F2, and PB1 are interdependent, and certain mutations previously used to ablate PB1-F2 production affected N40 accumulation. Removal of the PB1-F2 AUG upregulated N40 synthesis, while truncating PB1-F2 after codon 8 (with a concomitant M40I change in PB1) abolished N40 expression. A virus lacking both N40 and PB1-F2 replicated normally. However, viruses that did not express N40 but retained an intact PB1-F2 gene overexpressed PB1 early in infection and replicated slowly in tissue culture. Thus, the influenza A virus proteome includes a 12th primary translation product that (similarly to PB1-F2) is nonessential for virus viability but whose loss, in particular genetic backgrounds, is detrimental to virus replication.

Genome packaging in influenza A virus

“…The eight segments encode 12 identified polypeptides, only two of which are non-essential outside of the laboratory setting (Chen et al, 2001;Wise et al, 2009). As these two non-essential proteins are encoded by segment 2, along with the essential PB1 polymerase subunit, it follows that at least one copy of each of the eight vRNAs must be packaged for a single virion to be able to initiate a productive infection.…”

NLRX1 prevents mitochondrial induced apoptosis and enhances macrophage antiviral immunity by interacting with influenza virus PB1-F2 protein

“…IAV polymerase basic protein 1-frame 2 (PB1-F2) is a 75-to 90-aa-long accessory protein encoded by an ORF overlapping the PB1 ORF on segment 2 of the viral genome (19,20). PB1-F2 has been shown to modulate the induction of type I IFN (21)(22)(23); however, its main function is to induce apoptosis in host immune cells by targeting mitochondria via the basic amphipathic helix in its Significance Apoptosis refers to the ability of a cell to undergo programmed cell death under normal physiological conditions or in response to stress signals.…”

Influenza A virus utilizes a suboptimal Kozak sequence to fine-tune virus replication and host response

“…However, the difference could not be detected at later time points (8 and 12 h p.i.). Meanwhile, we also found that the expression of N40, a shorter form of PB1 protein that could be produced from the PB1 segment (Wise et al, 2009), was decreased in WSN-PB1(23A) virus infected cells. It has been reported that overexpression of N40 had a detrimental effect on virus replication in cell culture (Tauber et al, 2012).…”

“…In addition to the main IFN antagonist NS1, RdRp was also reported to be involved in host shut-off by targeting host Pol II transcription machinery (Graef et al, 2010;Vreede et al, 2010). Moreover, a short non-structural peptide PB1-F2, expressed from the PB1 segment of influenza A virus, was reported to be involved in modulating host antiviral status (Chen et al, 2001;Wise et al, 2009). Therefore, an understanding of the mechanisms by which influenza virus controls its RNA and protein expression, and the consequences of such regulation, is of great significance in order to fully understand influenza A virus replication strategies.…”