title: Contributions of the S2 spike ectodomain to attachment and host range of infectious bronchitis virus.
abstract: The spike protein is the major viral attachment protein of the avian coronavirus infectious bronchitis virus (IBV) and ultimately determines viral tropism. The S1 subunit of the spike is assumed to be required for virus attachment. However, we have previously shown that this domain of the embryo- and cell culture adapted Beaudette strain, in contrast to that of the virulent M41 strain, is not sufficient for binding to chicken trachea (Wickramasinghe et al., 2011). In the present study, we demonstrated that the lack of binding of Beaudette S1 was not due to absence of virus receptors on this tissue nor due to the production of S1 from mammalian cells, as S1 proteins expressed from chicken cells also lacked the ability to bind IBV-susceptible embryonic tissue. Subsequently, we addressed the contribution of the S2 subunit of the spike in IBV attachment. Recombinant IBV Beaudette spike ectodomains, comprising the entire S1 domain and the S2 ectodomain, were expressed and analyzed for binding to susceptible embryonic chorio-allantoic membrane (CAM) in our previously developed spike histochemistry assay. We observed that extension of the S1 domain with the S2 subunit of the Beaudette spike was sufficient to gain binding to CAM. A previously suggested heparin sulfate binding site in Beaudette S2 was not required for the observed binding to CAM, while sialic acids on the host tissues were essential for the attachment. To further elucidate the role of S2 the spike ectodomains of virulent IBV M41 and chimeras of M41 and Beaudette were analyzed for their binding to CAM, chicken trachea and mammalian cell lines. While the M41 spike ectodomain showed increased attachment to both CAM and chicken trachea, no binding to mammalian cells was observed. In contrast, Beaudette spike ectodomain had relatively weak ability to bind to chicken trachea, but displayed marked extended host range to mammalian cells. Binding patterns of chimeric spike ectodomains to these tissues and cells indicate that S2 subunits most likely do not contain an additional independent receptor-binding site. Rather, the interplay between S1 and S2 subunits of spikes from the same viral origin might finally determine the avidity and specificity of virus attachment and thus viral host range.
authors.affiliation: Pathology Division, Department of Pathobiology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584 CL Utrecht, The Netherlands.
authors.forename: N
authors.initials: N
authors.lastname: Promkuntod
authors.forename: I N Ambepitiya
authors.initials: IN
authors.lastname: Wickramasinghe
authors.forename: G
authors.initials: G
authors.lastname: de Vrieze
authors.forename: A
authors.initials: A
authors.lastname: Gröne
authors.forename: M H
authors.initials: MH
authors.lastname: Verheije
mesh_terms: D000818:Animals; D002460:Cell Line; D002642:Chick Embryo; D018352:Coronavirus Infections / Q000378:metabolism / Q000821:virology*; D058507:Host Specificity; D006801:Humans; D001351:Infectious bronchitis virus / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D017434:Protein Structure, Tertiary; D011991:Receptors, Virus / Q000378:metabolism; D064370:Spike Glycoprotein, Coronavirus / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D053585:Virus Attachment
chemical_list: D011991:Receptors, Virus; D064370:Spike Glycoprotein, Coronavirus
keywords: Attachment; Avian coronavirus; Beaudette; Infectious bronchitis virus; M41; Spike
references.citation: J Virol. 1999 Jan;73(1):638-49
references.pmid: 9847369
references.citation: J Virol. 2003 Aug;77(16):9084-9
references.pmid: 12885925
references.citation: Med Res Rev. 2002 Jan;22(1):1-25
references.pmid: 11746174
references.citation: J Virol. 2006 Nov;80(22):10909-18
references.pmid: 16956938
references.citation: Vet Microbiol. 2012 Jun 15;157(3-4):285-93
references.pmid: 22340967
references.citation: J Virol. 2003 Jan;77(2):841-50
references.pmid: 12502800
references.citation: Virology. 2004 Sep 1;326(2):288-98
references.pmid: 15302214
references.citation: J Gen Virol. 2006 May;87(Pt 5):1209-1216
references.pmid: 16603523
references.citation: Avian Pathol. 2009 Feb;38(1):41-5
references.pmid: 19156578
references.citation: J Virol. 2004 Jun;78(11):6048-54
references.pmid: 15141003
references.citation: PLoS One. 2011;6(8):e24352
references.pmid: 21912629
references.citation: J Virol. 1997 Dec;71(12):9499-507
references.pmid: 9371612
references.citation: Avian Pathol. 2011 Jun;40(3):223-35
references.pmid: 21711181
references.citation: J Virol. 1995 Sep;69(9):5535-43
references.pmid: 7636998
references.citation: J Gen Virol. 1972 Sep;16(3):423-7
references.pmid: 4627930
references.citation: J Virol. 2010 Apr;84(7):3134-46
references.pmid: 19906932
references.citation: Arch Virol. 1979;60(2):115-22
references.pmid: 226034
references.citation: Virology. 2012 Nov 25;433(2):421-30
references.pmid: 22995191
references.citation: Biochem Biophys Res Commun. 2005 Oct 21;336(2):417-23
references.pmid: 16137658
references.citation: J Virol. 2008 Feb;82(3):1414-24
references.pmid: 18032498
references.citation: Virology. 2004 Jul 1;324(2):510-24
references.pmid: 15207636
references.citation: Arteriosclerosis. 1989 Jan-Feb;9(1):21-32
references.pmid: 2463827
references.citation: Viruses. 2012 Jun;4(6):980-1010
references.pmid: 22816036
references.citation: Viruses. 2012 Jun;4(6):1011-33
references.pmid: 22816037
references.citation: J Virol. 2011 Sep;85(17):8903-12
references.pmid: 21697468
references.citation: J Virol. 1997 Dec;71(12):9024-31
references.pmid: 9371559
references.citation: J Virol. 2004 Sep;78(17):9073-83
references.pmid: 15308703
references.citation: J Virol. 1997 Mar;71(3):1946-55
references.pmid: 9032326
references.citation: Microbes Infect. 2008 Apr;10(4):367-73
references.pmid: 18396435
references.citation: Avian Dis. 2012 Dec;56(4):634-41
references.pmid: 23397833
references.citation: Virus Res. 1986 Feb;4(2):133-43
references.pmid: 3010595
references.citation: Adv Exp Med Biol. 1998;440:735-41
references.pmid: 9782352
references.citation: J Virol. 2004 Dec;78(24):13804-11
references.pmid: 15564488
references.citation: J Virol. 2005 Jun;79(12):7629-40
references.pmid: 15919915
references.citation: J Virol. 2009 Sep;83(17):8744-58
references.pmid: 19553314
references.citation: Res Vet Sci. 1976 May;20(3):348-9
references.pmid: 180581
references.citation: Arch Virol. 2012 Jul;157(7):1411-22
references.pmid: 22481600
references.citation: PLoS One. 2009 Jul 02;4(7):e6130
references.pmid: 19572016
references.citation: Avian Dis. 1985 Apr-Jun;29(2):546-51
references.pmid: 2992442
references.citation: Avian Dis. 2001 Apr-Jun;45(2):366-72
references.pmid: 11417816
references.citation: J Virol. 2005 Nov;79(22):14451-6
references.pmid: 16254381
references.citation: Biochemistry. 2006 Dec 26;45(51):15205-15
references.pmid: 17176042
references.citation: Viruses. 2012 Apr;4(4):557-80
references.pmid: 22590686
references.citation: Methods Mol Biol. 2008;454:109-17
references.pmid: 19057881
references.citation: Arch Gesamte Virusforsch. 1973;40(3):285-90
references.pmid: 4573193
references.citation: Avian Dis. 2007 Mar;51(1):45-51
references.pmid: 17461266
references.citation: J Virol. 2003 Aug;77(16):8801-11
references.pmid: 12885899