Epstein–Barr virus is associated with several human lymphomas and carcinomas, and its BARF1 oncogene encodes a protein that is thought to play an important role in carcinogenesis. A BARF1 recombinant adenovirus expression system, which led us to discover the macromolecular size of the cleaved and secreted form of the BARF1 protein in the native state and its mitogenic capacity on various cell lines in culture, was used further to investigate the structure and maturation of the BARF1 protein. We recently reported biophysical studies that showed dimer-based oligomerization of the BARF1 polypeptide. Here, new data are presented that confirm post-translational modifications predicted from the BARF1 sequence: phosphorylation on serine and threonine, and N- and O-glycosylation. The N- and O-glycans were partially characterized and it was demonstrated that both modifications are required for active secretion of the BARF1 protein via the classical pathway.
CarpentierM.,
MorelleW.,
CoddevilleB.,
PonsA.,
MassonM.,
MazurierJ.,
LegrandD.2005; Nucleolin undergoes partial N - and O -glycosylations in the extranuclear cell compartment. Biochemistry 44:5804–5815[CrossRef]
CohenJ. I.,
LekstromK.1999; Epstein–Barr virus BARF1 protein is dispensable for B-cell transformation and inhibits alpha interferon secretion from mononuclear cells. J Virol 73:7627–7632
de Turenne-TessierM.,
JolicoeurP.,
OokaT.1997; Expression of the protein encoded by Epstein–Barr virus (EBV) BARF1 open reading frame from a recombinant adenovirus system. Virus Res 52:73–85[CrossRef]
de Turenne-TessierM.,
JolicoeurP.,
MiddeldorpJ. M.,
OokaT.2005; Expression and analysis of the Epstein–Barr virus BARF1-encoded protein from a tetracycline-regulatable adenovirus system. Virus Res 109:9–18[CrossRef]
DecaussinG.,
Sbih-LammaliF.,
de Turenne-TessierM.,
BouguermouhA.,
OokaT.2000; Expression of BARF1 gene encoded by Epstein–Barr virus in nasopharyngeal carcinoma biopsies. Cancer Res 60:5584–5588
EvansA. G.,
MoormanN. J.,
WillerD. O.,
SpeckS. H.2006; The M4 gene of γ HV68 encodes a secreted glycoprotein and is required for the efficient establishment of splenic latency. Virology 344:520–531[CrossRef]
MaleyF.,
TrimbleR. B.,
TarentinoA. L.,
PlummerT. H., Jr.
1989; Characterization of glycoproteins and their associated oligosaccharides through the use of endoglycosidases. Anal Biochem 180:195–204[CrossRef]
MardassiH.,
GoninP.,
GagnonC. A.,
MassieB.,
DeaS.1998; A subset of porcine reproductive and respiratory syndrome virus GP3 glycoprotein is released into the culture medium of cells as a non-virion-associated and membrane-free (soluble) form. J Virol 72:6298–6306
MeadsM. B.,
MedveczkyP. G.2004; Kaposi's sarcoma-associated herpesvirus encoded viral interleukin-6 is secreted and modified differently than human interleukin-6: evidence for a unique autocrine signaling mechanism. J Biol Chem 279:51793–51803[CrossRef]
MitraN.,
SinhaS.,
RamyaT. N. C.,
SuroliaA.2006; N -linked oligosaccharides as outfitters for glycoprotein folding, form and function. Trends Biochem Sci 31:156–163[CrossRef]
NaganoM.,
StübigerG.,
MarchettiM.,
GmeinerG.,
AllmaierG.,
ReichelC.2005; Detection of isoforms of recombinant human erythropoietin by various plant lectins after isoelectric focusing. Electrophoresis 26:1633–1645[CrossRef]
NgA.,
TscharkeD. C.,
ReadingP. C.,
SmithG. L.2001; The vaccinia virus A41L protein is a soluble 30 kDa glycoprotein that affects virus virulence. J Gen Virol 82:2095–2105
OokaT.2005; Biological role of the BARF1 gene encoded by Epstein–Barr virus. In Epstein–Barr Virus pp 613–630 Edited by
RobertsonE. S.
Norfolk, UK: Caister Academic Press;
PagerC. T.,
WurthM. A.,
DutchR. E.2004; Subcellular localization and calcium and pH requirements for proteolytic processing of the Hendra virus fusion protein. J Virol 78:9154–9163[CrossRef]
RessingM. E.,
van LeeuwenD.,
VerreckF. A.,
KeatingS.,
GomezR.,
FrankenK. L.,
OttenhoffT. H.,
SpriggsM.,
SchumacherT. N.other authors2005; Epstein–Barr virus gp42 is posttranslationally modified to produce soluble gp42 that mediates HLA class II immune evasion. J Virol 79:841–852[CrossRef]
SalazarC.,
HöferT.2007; Versatile regulation of multisite protein phosphorylation by the order of phosphate processing and protein-protein interactions. FEBS J 274:1046–1061[CrossRef]
TannerJ. E.,
WeiM. X.,
AlfieriC.,
AhmadA.,
TaylorP.,
OokaT.,
MenezesJ.1997; Antibody and antibody-dependent cellular cytotoxicity responses against the Bam H1 A rightward open-reading frame-1 protein of Epstein–Barr virus (EBV) in EBV-associated disorders. J Infect Dis 175:38–46[CrossRef]
Van den SteenP.,
RuddP. M.,
DwekR. A.,
OpdenakkerG.1998; Concepts and principles of O -linked glycosylation. Crit Rev Biochem Mol Biol 33:151–208[CrossRef]
Wahl-JensenV.,
KurzS. K.,
HazeltonP. R.,
SchnittlerH. J.,
StroherU.,
BurtonD. R.,
FeldmannH.2005; Role of Ebola virus secreted glycoproteins and virus-like particles in activation of human macrophages. J Virol 79:2413–2419[CrossRef]
WangL.,
TamJ. P.,
LiuD. X.2006a; Biochemical and functional characterization of Epstein–Barr virus-encoded BARF1 protein: interaction with human hTid1 protein facilitates its maturation and secretion. Oncogene 25:4320–4331[CrossRef]
WangQ.,
TsaoS. W.,
OokaT.,
NichollsJ. M.,
CheungH. W.,
FuS.,
WongY. C.,
WangX.2006b; Anti-apoptotic role of BARF1 in gastric cancer cells. Cancer Lett 238:90–103[CrossRef]
zur HausenA.,
BrinkA. A.,
CraanenM. E.,
MiddeldorpJ. M.,
MeijerC. J.,
van den BruleA. J.2000; Unique transcription pattern of Epstein–Barr virus (EBV) in EBV-carrying gastric adenocarcinomas: expression of the transforming BARF1 gene. Cancer Res 60:2745–2748