signature=0656384d75135bfca37995b17607e69c,Adaptation of avian influenza A virus polymerase in mamma...

Adaptation of Avian In?uenza A Virus Polymerase in Mammals To Overcome the Host Species Barrier

Benjamin M?nz,*Martin Schwemmle,Linda Brunotte

Department of Virology,Institute for Medical Microbiology and Hygiene,University of Freiburg,Freiburg,Germany

Avian in?uenza A viruses,such as the highly pathogenic avian H5N1viruses,sporadically enter the human population but often do not transmit between individuals.In rare cases,however,they establish a new lineage in humans.In addition to well-charac-terized barriers to cell entry,one major hurdle which avian viruses must overcome is their poor polymerase activity in human cells.There is compelling evidence that these viruses overcome this obstacle by acquiring adaptive mutations in the polymerase subunits PB1,PB2,and PA and the nucleoprotein(NP)as well as in the novel polymerase cofactor nuclear export protein(NEP). Recent?ndings suggest that synthesis of the viral genome may represent the major defect of avian polymerases in human cells. While the precise mechanisms remain to be unveiled,it appears that a broad spectrum of polymerase adaptive mutations can act collectively to overcome this defect.Thus,identi?cation and monitoring of emerging adaptive mutations that further increase polymerase activity in human cells are critical to estimate the pandemic potential of avian viruses.

A lthough the natural reservoirs of in?uenza A viruses lacking

adaptive mutations that increase polymerase activity in mam-malian cells(1–53)are wild birds(54),mammals are frequently infected with in?uenza viruses of avian origin.These zoonotic transmissions can cause severe disease in different mammals,in-cluding cats,dogs,horses,pigs,and humans(55–57),due to the lack of preexisting immunity in these species to the new in?uenza virus strain(Fig.1).Fortunately,most of these infections are so-called dead-end infections and are not further transmitted within the new species due to several barriers.However,on rare occasions in?uenza A viruses can indeed break the species barrier and estab-lish an entirely new virus lineage in a mammalian species,as ex-empli?ed by the human pandemic of1918(58),the Eurasian clas-sical swine in?uenza virus lineage(59),H3N8in?uenza viruses in horses(56),and possibly H17N10-like in?uenza viruses in bats (60).Although16hemagglutinin(HA)and9neuraminidase (NA)subtypes have been identi?ed in wild birds,human infection has been documented only for H1,H2,H3,H5,H7,and H9(61–64)and only H1,H2,and H3have been stably introduced into the human population(56,65,66).In the majority of cases,human pandemics were a result of genetic reassortment events whereby the circulating human virus strains acquired one or more gene segments from avian or swine sources(54).However,since hu-mans in certain regions of the globe are constantly exposed to H5N1in?uenza viruses,there exists a serious concern that this subtype might acquire mutations to stably cross the species barrier and start a new pandemic(67,68).

REPLICATION OF INFLUENZA A VIRUSES

In?uenza viruses belong to the family of Orthomyxoviridae,pos-sessing a single-stranded negative-sense RNA genome that is com-prised of eight segments(69).The ends of each genome segment are short complementary elements that form the viral promoter and are recognized by the viral RNA-dependent RNA polymerase, which is composed of the three subunits PB2,PB1,and PA(70–74).Together,the viral polymerase,the nucleoprotein NP,and the viral RNA genome form the ribonucleoprotein(RNP)complex, which is responsible for viral mRNA synthesis and genome repli-cation.Upon infection,viral RNPs(vRNPs)are released into the cytoplasm(75)and transported into the nucleus,where the RNPs perform all of their enzymatic functions(76,77).First,primary transcription is initiated and viral mRNA is synthesized from the viral genome(vRNA)(Fig.2).vRNA also serves as a template for synthesis of a full-length copy(cRNA),which serves as a template for subsequent synthesis of new viral genomes(78).Viral mRNA synthesis is primed using a cellular capped pre-mRNA which is bound by the PB2subunit(30,79)and cleaved by the endonu-cleolytic domain of the polymerase subunit PA(39,80).Synthesis of both c-and vRNA is apparently primer independent(81–83). Recent evidence suggests that mRNA is synthesized by viral poly-merase complexes which are resident on vRNPs(in“cis”),whereas vRNA is synthesized from the cRNA template using soluble viral polymerase complexes(in“trans”)(84).However,the mecha-nism of cRNA synthesis and the regulation of the switch between transcription and genome replication remain largely unknown (85).The viral nucleoprotein(NP)is one major candidate to reg-ulate the switch between transcription and replication(86),pre-sumably by direct interaction with the viral polymerase(87),al-though this role of NP has been questioned more recently(88).

Interestingly,in addition to the viral nucleoprotein,the nu-clear export protein NEP(NS2)has been shown to be involved in the regulation of polymerase activity(11,89,90).In contrast to NP,it is not an essential component of the RNP complex,as it is not required for transcriptional activity in a polymerase reconsti-tution assay(Fig.3).In this system,the effect of NEP on viral RNA synthesis was shown to be concentration dependent.While very high NEP concentrations completely abrogated polymerase activ-ity(11),recent data demonstrate that small amounts of NEP stim-Published ahead of print24April2013

Address correspondence to Martin Schwemmle,

[e-mail].

*Present address:Benjamin M?nz,Department of Viroscience,Erasmus MC,

Rotterdam,The Netherlands.

Copyright?2013,American Society for Microbiology.All Rights Reserved.

doi:10.1128/JVI.00980-13

MINIREVIEW

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