Epstein–Barr virus-encoded LMP-1 and CD40 mediate IL-6 production in epithelial cell via an NF-kB pathway involving TNF receptor-associated factors

Harvard University, Cambridge, Massachusetts, United States
Oncogene (Impact Factor: 8.46). 07/1997; 14(24):2899-916. DOI: 10.1038/sj.onc.1201258
Source: PubMed

ABSTRACT Expression of the Epstein-Barr virus (EBV) transforming LMP1 in B cells activates the transcription factor NF-kappaB and induces phenotypic changes through two distinct domains in the cytoplasmic C-terminus of the protein. The aa 187-231 domain of LMP1, which is important for growth transformation, binds tumour necrosis factor (TNF) receptor associated factor (TRAF) 1 and TRAF3 and this interaction mediates subsequent signalling events. The TRAFs also associate with CD40, a member of the TNFR family, which upon ligation activates NF-kappaB and induces phenotypic changes similar to those mediated by LMP1. This study demonstrates that LMP1 expression in carcinoma cell lines and SV40-transformed keratinocytes results in induction of the pleiotropic cytokine interleukin 6 (IL6), an effect which is also observed upon CD40 ligation. The mechanism by which either LMP1 expression or CD40 ligation induces IL6 production was found to be NF-kappaB-dependent. Mutational analysis identified domains in the C-terminus of LMP1 which are important for NF-kappaB activation and IL6 secretion. LMP1 and CD40 share a common PxQxT core TRAF binding motif and mutations in or adjacent to this sequence impaired the ability of LMP1 or CD40 to induce NF-kappaB activation and IL6 secretion. The importance of TRAF interactions in mediating these effects was confirmed using dominant negative TRAF2 and TRAF3 mutants which also identified differences in the signalling events mediated by the two NF-kappaB activating domains of LMP1. A20, an anti-apoptotic protein which interacts with TRAF2 and blocks CD40-mediated NF-kappaB activity, also blocked NF-kappaB and IL6 secretion in LMP1-transfected epithelial cells. These results suggest that LMP1 regulates IL6 production in epithelial cells in a manner similar to CD40 ligation and implicate TRAFs as common mediators in the transduction of signals generated via the CD40 and LMP1 pathways. As a role for IL6 in regulating epithelial cell growth has previously been suggested, the control of IL6 secretion via the CD40 and LMP1 pathways may have implications for the growth of both normal and transformed epithelial cells.

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Available from: Lawrence S Young, Jun 11, 2014
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    • "The STAT3 activation by IL-6 is well-documented to support growth, survival and metastasis of human cancer cells [14], [16]. In vitro, this enhanced cellular response to IL-6-induced STAT3 activation may be beneficial to immortalized NPE cells to overcome the stress-induced growth inhibition commonly associated with EBV infection [23], [25]. In our immortalized NPE cell models, overexpression of IL-6R was found to be primarily responsible for the enhanced response to IL6-activation of STAT3 signaling (Fig 2). "
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    ABSTRACT: Nasopharyngeal carcinoma (NPC) is etiologically associated with Epstein-Barr virus (EBV) infection. However, the exact role of EBV in NPC pathogenesis remains elusive. Activation of signal transducer and activator of transcription 3 (STAT3) is common in human cancers including NPC and plays an important role in the pathogenesis and progression of human cancers. Interleukin-6 (IL-6), a major inflammatory cytokine, is a potent activator of STAT3. In this study, we report that EBVinfected immortalized nasopharyngeal epithelial (NPE) cells often acquire an enhanced response to IL-6-induced STAT3 activation to promote their growth and invasive properties. Interestingly, this enhanced IL-6/STAT3 response was mediated by overexpression of IL-6 receptor (IL-6R). Furthermore, IL-6R overexpression enhanced IL-6-induced STAT3 activation in uninfected immortalized NPE cells in vitro, and promoted growth and tumorigenicity of EBV-positive NPC cell line (C666-1) in vivo. Moreover, it is shown for the first time that IL-6R was overexpressed in clinical specimens of NPC. IL-6 expression could also be strongly detected in the stromal cells of NPC and a higher circulating level of IL-6 was found in the sera of advance-staged NPC patients compared to the control subjects. Therefore, IL-6R overexpression, coupled with enhanced IL-6/STAT3 signaling may facilitate the malignant transformation of EBV-infected premalignant NPE cells into cancer cells, and enhance malignant properties of NPC cells.
    PLoS ONE 05/2013; 8(5):e62284. DOI:10.1371/journal.pone.0062284 · 3.23 Impact Factor
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    • "The major oncogene of EBV, LMP1, is a functional CD40 mimic. Interactions of cellular tumor necrosis factor (TNF) receptor-associated factor (TRAF) adaptor proteins, including TRAF1 [7], [8] and TRAF3 [7], [9], with the LMP1 C-terminal tail signaling domains, carboxy-terminal activating region 1 and 2 (CTAR1 or CTAR2), initiate signal transduction through a variety of pathways including the p38 [10], [11], Erk [12], [13], and JNK [14], [15] MAPK and NF-κB [16], [17] pathways. "
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    ABSTRACT: The B lymphotrophic γ-herpesvirus EBV is associated with a variety of lymphoid- and epithelial-derived malignancies, including B cell lymphomas in immunocompromised and immunosuppressed individuals. The primary oncogene of EBV, latent membrane protein 1 (LMP1), activates the PI3K/Akt pathway to induce the autocrine growth factor, IL-10, in EBV-infected B cells, but the mechanisms underlying PI3K activation remain incompletely understood. Using small molecule inhibition and siRNA strategies in human B cell lines expressing a chimeric, signaling-inducible LMP1 protein, nerve growth factor receptor (NGFR)-LMP1, we show that NGFR-LMP1 utilizes Syk to activate PI3K/Akt signaling and induce IL-10 production. NGFR-LMP1 signaling induces phosphorylation of BLNK, a marker of Syk activation. Whereas Src kinases are often required for Syk activation, we show here that PI3K/Akt activation and autocrine IL-10 production by NGFR-LMP1 involves the Src family kinase Fyn. Finally, we demonstrate that NGFR-LMP1 induces phosphorylation of c-Cbl in a Syk- and Fyn-dependent fashion. Our results indicate that the EBV protein LMP1, which lacks the canonical ITAM required for Syk activation, can nevertheless activate Syk, and the Src kinase Fyn, resulting in downstream c-Cbl and PI3K/Akt activation. Fyn, Syk, and PI3K/Akt antagonists thus may present potential new therapeutic strategies that target the oncogene LMP1 for treatment of EBV+ B cell lymphomas.
    PLoS ONE 08/2012; 7(8):e42610. DOI:10.1371/journal.pone.0042610 · 3.23 Impact Factor
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    • "Latent membrane protein 1 acts as a constitutively active member of the tumour necrosis factor receptor family (CD40) (Uchida et al, 1999), activating a multitude of intracellular signalling pathways in a ligand-independent manner. Particularly, the STAT3 pathway can be strongly activated by LMP1 via JAK3 kinase (Eliopoulos et al, 1999; Gires et al, 1999; Chen et al, 2003) or an increase of IL6 secretion (Eliopoulos et al, 1997; Hirano et al, 2000). Signal transducer and activator of transcription 3, in turn, up-regulates c-myc (Bromberg et al, 1999; Bowman et al, 2001), which has been shown to down-regulate HLA class I APM expression in human melanoma cell lines (Versteeg et al, 1988; Blom et al, 1997), several carcinoma cell lines (Ottesen et al, 1990; Belldegrun et al, 1993) and in B cells (Staege et al, 2002). "
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    ABSTRACT: Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1) up-regulates the human leukocyte antigen (HLA) class I antigen presentation machinery (APM). This appears counterintuitive with immune evasion in EBV-associated tumours like nasopharyngeal carcinoma (NPC). Latent membrane protein 1-transfected epithelial cell lines were used as a model system to study the impact of LMP1 and c-Myc on HLA class I components. The expression of components of the HLA class I APM, c-Myc and Ki-67 was analysed in LMP1+ and LMP1- NPC by immunohistochemistry. In epithelial cells, LMP1 up-regulated HLA class I APM. This effect could be counteracted by c-Myc, which itself was up-regulated by LMP1 apparently through IL6 induction and Jak3/STAT3 activation. Studies of NPC biopsies revealed down-regulation of HLA class I APM expression. No difference was observed between LMP1+ and LMP1- NPC. However, expression of Ki-67 and c-Myc were up-regulated in LMP1+ tumours. These findings raise the possibility that c-Myc activation in NPC might antagonise the effect of LMP1 on HLA class I expression thus contributing to immune escape of tumour cells.
    British Journal of Cancer 05/2012; 106(12):1980-8. DOI:10.1038/bjc.2012.197 · 4.84 Impact Factor
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