Porcine sapovirus (PSaV) of the family Caliciviridae, is the only member of the genus Sapovirus with cell culture and reverse genetics systems. When combined with the piglet model, these approaches provide a system to understand the molecular basis of sapovirus pathogenesis. The replication of PSaV in cell culture is, however, restricted, displaying an absolute requirement for bile acids and producing lower levels of infectious virus than other caliciviruses. The effect of bile acids has previously been linked to a reduction in the signal transducer and activator of transcription (STAT1)-mediated signalling pathway. In the current study, we observed that even in the presence of bile acids, PSaV replication in cell culture was restricted by soluble factors produced from infected cells. This effect was at least partially due to secreted IFN because treatment of cells with recombinant porcine IFN-β resulted in significantly reduced viral replication. Moreover, IFN-mediated signalling pathways (IFN, STAT1 and the 2′,5′-oligoadenylate synthetase) were activated during PSaV infection. Characterization of PSaV growth in cell lines deficient in their ability to induce or respond to IFN showed a 100–150-fold increase in infectious virus production, indicating that the primary role of bile acids was not the inactivation of the innate immune response. Furthermore, the use of IFN-deficient cell lines enabled more efficient recovery of PSaV from cDNA constructs. Overall, the highly efficient cell culture and reverse genetics system established here for PSaV highlighted the key role of the innate immune response in the restriction of PSaV infection and should greatly facilitate further molecular studies on sapovirus host–cell interactions.
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AndrusL.,
MarukianS.,
JonesC. T.,
CataneseM. T.,
SheahanT. P.,
SchogginsJ. W.,
BarryW. T.,
DustinL. B.,
TrehanK.other authors2011; Expression of paramyxovirus V proteins promotes replication and spread of hepatitis C virus in cultures of primary human fetal liver cells. Hepatology 54:1901–1912 [View Article][PubMed]
ApelbaumA.,
YardenG.,
WarszawskiS.,
HarariD.,
SchreiberG.2013; Type I interferons induce apoptosis by balancing cFLIP and caspase-8 independent of death ligands. Mol Cell Biol 33:800–814 [View Article][PubMed]
BekiszJ.,
BaronS.,
BalinskyC.,
MorrowA.,
ZoonK. C.2010; Antiproliferative properties of type I and type II interferon. Pharmaceuticals (Basel) 3:994–1015 [View Article][PubMed]
BlantonL. H.,
AdamsS. M.,
BeardR. S.,
WeiG.,
BulensS. N.,
WiddowsonM.-A.,
GlassR. I.,
MonroeS. S.2006; Molecular and epidemiologic trends of caliciviruses associated with outbreaks of acute gastroenteritis in the United States, 2000–2004. J Infect Dis 193:413–421 [View Article][PubMed]
ChangK.-O.,
GeorgeD. W.2007; Bile acids promote the expression of hepatitis C virus in replicon-harboring cells. J Virol 81:9633–9640 [View Article][PubMed]
ChangK.-O.,
SosnovtsevS. V.,
BelliotG.,
KimY.,
SaifL. J.,
GreenK. Y.2004; Bile acids are essential for porcine enteric calicivirus replication in association with down-regulation of signal transducer and activator of transcription 1. Proc Natl Acad Sci U S A 101:8733–8738 [View Article][PubMed]
ChangotraH.,
JiaY.,
MooreT. N.,
LiuG.,
KahanS. M.,
SosnovtsevS. V.,
KarstS. M.2009; Type I and type II interferons inhibit the translation of murine norovirus proteins. J Virol 83:5683–5692 [View Article][PubMed]
ChaudhryY.,
SkinnerM. A.,
GoodfellowI. G.2007; Recovery of genetically defined murine norovirus in tissue culture by using a fowlpox virus expressing T7 RNA polymerase. J Gen Virol 88:2091–2100 [View Article][PubMed]
ChildsK.,
StockN.,
RossC.,
AndrejevaJ.,
HiltonL.,
SkinnerM.,
RandallR.,
GoodbournS.2007; mda-5, but not RIG-I, is a common target for paramyxovirus V proteins. Virology 359:190–200 [View Article][PubMed]
HaymanA.,
ComelyS.,
LackenbyA.,
HartgrovesL. C. S.,
GoodbournS.,
McCauleyJ. W.,
BarclayW. S.2007; NS1 proteins of avian influenza A viruses can act as antagonists of the human alpha/beta interferon response. J Virol 81:2318–2327 [View Article][PubMed]
HiltonL.,
MoganeradjK.,
ZhangG.,
ChenY.-H.,
RandallR. E.,
McCauleyJ. W.,
GoodbournS.2006; The NPro product of bovine viral diarrhea virus inhibits DNA binding by interferon regulatory factor 3 and targets it for proteasomal degradation. J Virol 80:11723–11732 [View Article][PubMed]
McFaddenN.,
BaileyD.,
CarraraG.,
BensonA.,
ChaudhryY.,
ShortlandA.,
HeeneyJ.,
YarovinskyF.,
SimmondsP.other authors2011; Norovirus regulation of the innate immune response and apoptosis occurs via the product of the alternative open reading frame 4. PLoS Pathog 7:e1002413 [View Article][PubMed]
NakamuraK.,
SagaY.,
IwaiM.,
ObaraM.,
HorimotoE.,
HasegawaS.,
KurataT.,
OkumuraH.,
NagoshiM.,
TakizawaT.2010; Frequent detection of noroviruses and sapoviruses in swine and high genetic diversity of porcine sapovirus in Japan during Fiscal Year 2008. J Clin Microbiol 48:1215–1222 [View Article][PubMed]
ReuterG.,
Zimsek-MijovskiJ.,
Poljsak-PrijateljM.,
Di BartoloI.,
RuggeriF. M.,
KantalaT.,
MaunulaL.,
KissI.,
KecskemétiS.other authors2010; Incidence, diversity, and molecular epidemiology of sapoviruses in swine across Europe. J Clin Microbiol 48:363–368 [View Article][PubMed]
SherwoodV.,
BurgertH.-G.,
ChenY.-H.,
SangheraS.,
KatafigiotisS.,
RandallR. E.,
ConnertonI.,
MellitsK. H.2007; Improved growth of enteric adenovirus type 40 in a modified cell line that can no longer respond to interferon stimulation. J Gen Virol 88:71–76 [View Article][PubMed]
ShivannaV.,
KimY.,
ChangK.-O.2014; The crucial role of bile acids in the entry of porcine enteric calicivirus. Virology 456–457:268–278 [View Article][PubMed]
ThiryE.,
PoelW. H.,
MauroyA.,
ThysC.,
HoningR. H.-V.2012; Development and application of a SYBR green RT-PCR for first line screening and quantification of porcine sapovirus infection. BMC Vet Res 8:193[CrossRef]
WangQ.-H.,
SouzaM.,
FunkJ. A.,
ZhangW.,
SaifL. J.2006; Prevalence of noroviruses and sapoviruses in swine of various ages determined by reverse transcription-PCR and microwell hybridization assays. J Clin Microbiol 44:2057–2062 [View Article][PubMed]
WobusC. E.,
ThackrayL. B.,
VirginH. W.IV2006; Murine norovirus: a model system to study norovirus biology and pathogenesis. J Virol 80:5104–5112 [View Article][PubMed]
YoungD. F.,
AndrejevaL.,
LivingstoneA.,
GoodbournS.,
LambR. A.,
CollinsP. L.,
ElliottR. M.,
RandallR. E.2003; Virus replication in engineered human cells that do not respond to interferons. J Virol 77:2174–2181 [View Article][PubMed]
YunusM. A.,
ChungL. M. W.,
ChaudhryY.,
BaileyD.,
GoodfellowI.2010; Development of an optimized RNA-based murine norovirus reverse genetics system. J Virol Methods 169:112–118 [View Article][PubMed]