Role of the TRAF binding site and NF-kappaB activation in Epstein-Barr virus latent membrane protein 1-induced cell gene expression.
In this study, we investigated the induction of cellular gene expression by the Epstein-Barr Virus (EBV) latent membrane protein 1 (LMP1). Previously, LMP1 was shown to induce the expression of ICAM-1, LFA-3, CD40, and EBI3 in EBV-negative Burkitt lymphoma (BL) cells and of the epidermal growth factor receptor (EGF-R) in epithelial cells. We now show that LMP1 expression also increased Fas and tumor necrosis factor receptor-associated factor 1 (TRAF1) in BL cells. LMP1 mediates NF-kappaB activation via two independent domains located in its C-terminal cytoplasmic tail, a TRAF-interacting site that associates with TRAF1, -2, -3, and -5 through a PXQXT/S core motif and a TRADD-interacting site. In EBV-transformed B cells or transiently transfected BL cells, significant amounts of TRAF1, -2, -3, and -5 are associated with LMP1. In epithelial cells, very little TRAF1 is expressed, and only TRAF2, -3, and -5, are significantly complexed with LMP1. The importance of TRAF binding to the PXQXT/S motif in LMP1-mediated gene induction was studied by using an LMP1 mutant that contains alanine point mutations in this motif and fails to associate with TRAFs. This mutant, LMP1(P204A/Q206A), induced 60% of wild-type LMP1 NF-kappaB activation and had approximately 60% of wild-type LMP1 effect on Fas, ICAM-1, CD40, and LFA-3 induction. In contrast, LMP1(P204A/Q206A) was substantially more impaired in TRAF1, EBI3, and EGF-R induction. Thus, TRAF binding to the PXQXT/S motif has a nonessential role in up-regulating Fas, ICAM-1, CD40, and LFA-3 expression and a critical role in up-regulating TRAF1, EBI3, and EGF-R expression. Further, D1 LMP1, an LMP1 mutant that does not aggregate failed to induce TRAF1, EBI3, Fas, ICAM-1, CD40, and LFA-3 expression confirming the essential role for aggregation in LMP1 signaling. Overexpression of a dominant form of IkappaBalpha blocked LMP1-mediated TRAF1, EBI3, Fas, ICAM-1, CD40, and LFA-3 up-regulation, indicating that NF-kappaB is an important component of LMP1-mediated gene induction from both the TRAF- and TRADD-interacting sites.
Devergne
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O
INSERM U131 and Institut Paris-Sud sur les Cytokines, 92140 Clamart, France.
Cahir McFarland
E D
ED
Mosialos
G
G
Izumi
K M
KM
Ware
C F
CF
Kieff
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E
D001665:Binding Sites; D002461:Cell Line, Transformed; D066246:ErbB Receptors / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D004854:Herpesvirus 4, Human / Q000378:metabolism*; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics; D016328:NF-kappa B / Q000378:metabolism*; D011485:Protein Binding; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism*; D015533:Transcriptional Activation; D015854:Up-Regulation; D014763:Viral Matrix Proteins / Q000378:metabolism*
C037662:EBV-associated membrane antigen, Epstein-Barr virus; D016328:NF-kappa B; D018124:Receptors, Tumor Necrosis Factor; D014763:Viral Matrix Proteins; D018799:Intercellular Adhesion Molecule-1; D066246:ErbB Receptors