Molecular pharmacological phenotyping of EBI2. An orphan seven-transmembrane receptor with constitutive activity.
Epstein-Barr virus (EBV)-induced receptor 2 (EBI2) is an orphan seven-transmembrane (7TM) receptor originally identified as the most up-regulated gene (>200-fold) in EBV-infected cells. Here we show that EBI2 signals with constitutive activity through Galpha(i) as determined by a receptor-mediated inhibition of forskolin-induced cAMP production and an induction of the serum response element-driven transcriptional activity in a pertussis toxin-sensitive manner. Galpha(s) and Galpha(q) were not activated constitutively as determined by the lack of cAMP production, the lack of inositol phosphate turnover, and the lack of activities of the transcription factors: cAMP response element-binding protein and nuclear factor-kappaB. Immunohistochemistry and confocal microscopy of FLAG- and green fluorescent protein-tagged EBI2 revealed cell-surface expression. A putative N-terminal truncated version of EBI2, delta4-EBI2, showed similar expression and signaling through Galpha(i) as full-length EBI2. By using a 32P-labeled EBI2 probe we found a very high expression in lymphoid tissue (spleen and lymph node) and peripheral blood mononuclear cells and a high expression in lung tissue. Real-time PCR of EBV-infected cells showed high expression of EBI2 during latent and lytic infection, in contrast to the EBV-encoded 7TM receptor BILF1, which was induced during lytic infection. EBI2 clustered with the orphan GPR18 by alignment analysis as well as by close proximity in the chromosomal region 13q32.3. Based on the constitutive signaling and cellular expression pattern of EBI2, it is suggested that it may function in conjunction with BILF1 in the reprogramming of the cell during EBV infection.
Rosenkilde
Mette M
MM
Laboratory for Molecular Pharmacology, Department of Pharmacology, University of Copenhagen, 2200 Copenhagen N, Copenhagen, Denmark. Electronic address: rosenkilde@molpharm.dk.
Benned-Jensen
Tau
T
Laboratory for Molecular Pharmacology, Department of Pharmacology, University of Copenhagen, 2200 Copenhagen N, Copenhagen, Denmark.
Andersen
Helene
H
Clinical Research Unit, Copenhagen University Hospital, 2650 Hvidovre, Denmark.
Holst
Peter J
PJ
Institute for Medical Microbiology and Immunology, The Panum Institute, Building 18.6, Blegdamsvej 3, 2200 Copenhagen N, Copenhagen, Denmark.
Kledal
Thomas N
TN
Clinical Research Unit, Copenhagen University Hospital, 2650 Hvidovre, Denmark.
Lüttichau
Hans R
HR
Laboratory for Molecular Pharmacology, Department of Pharmacology, University of Copenhagen, 2200 Copenhagen N, Copenhagen, Denmark; Department for Infectious Diseases, Copenhagen University Hospital, 2200 Copenhagen, Denmark.
Larsen
Jørgen K
JK
Finsen Laboratory, The Finsen Center, Copenhagen University Hospital, 2200 Copenhagen, Denmark.
Christensen
Jan P
JP
Institute for Medical Microbiology and Immunology, The Panum Institute, Building 18.6, Blegdamsvej 3, 2200 Copenhagen N, Copenhagen, Denmark.
Schwartz
Thue W
TW
Laboratory for Molecular Pharmacology, Department of Pharmacology, University of Copenhagen, 2200 Copenhagen N, Copenhagen, Denmark.
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D019204:GTP-Binding Proteins / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D007694:Killer Cells, Natural / Q000378:metabolism; D008214:Lymphocytes / Q000378:metabolism; D009000:Monocytes / Q000378:metabolism; D020033:Protein Isoforms; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction
C107515:GPR18 protein, human; C080105:GPR183 protein, human; D020033:Protein Isoforms; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D019204:GTP-Binding Proteins