title: Epstein-Barr virus-encoded BILF1 is a constitutively active G protein-coupled receptor.
abstract: Both beta- and gammaherpesviruses encode G protein-coupled receptors (GPCRs) with unique pharmacological phenotypes and important biological functions. An example is ORF74, the gamma2-herpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded GPCR, which is highly constitutively active and considered the key oncogene in Kaposi's sarcoma pathogenesis. In contrast, the current annotation of the Epstein-Barr virus (EBV) genome does not reveal any GPCR homolog encoded by this human oncogenic gamma1-herpesvirus. However, by employing bioinformatics, we recognized that the previously established EBV open reading frame BILF1 indeed encodes a GPCR. Additionally, BILF1 is a member of a new family of related GPCRs exclusively encoded by gamma1-herpesviruses. Expression of hemagglutinin-tagged BILF1 in the HEK293 epithelial cell line revealed that BILF1 is expressed as an approximately 50-kDa glycosylated protein. Immunocytochemistry and confocal microscopy revealed that BILF1 localizes predominantly to the plasma membrane, similar to the localization of KSHV ORF74. Using chimeric G proteins, we found that human and rhesus EBV-encoded BILF1 are highly potent constitutively active receptors, activating Galphai. Furthermore, BILF1 is able to inhibit forskolin-triggered CREB activation via stimulation of endogenous G proteins in a pertussis toxin-sensitive manner, verifying that BILF1 signals constitutively through Galphai. We suggest that EBV may use BILF1 to regulate Galphai-activated pathways during viral lytic replication, thereby affecting disease progression.
authors.affiliation: Clinical Research Unit, Copenhagen University Hospital, Hvidovre, Denmark.
authors.forename: Sarah J
authors.initials: SJ
authors.lastname: Paulsen
authors.forename: Mette M
authors.initials: MM
authors.lastname: Rosenkilde
authors.forename: Jesper
authors.initials: J
authors.lastname: Eugen-Olsen
authors.forename: Thomas N
authors.initials: TN
authors.lastname: Kledal
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D006031:Glycosylation; D004854:Herpesvirus 4, Human / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D006801:Humans; D007150:Immunohistochemistry; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D043562:Receptors, G-Protein-Coupled; D014764:Viral Proteins; D019206:GTP-Binding Protein alpha Subunits, Gi-Go