Ubiquitination of the Epstein–Barr virus-encoded latent membrane protein 1 depends on the integrity of the TRAF binding site

Rothenberger, Sylvia; Burns, Kimberly; Rousseaux, Marga; Tschopp, Jürg; Bron, Claude

doi:10.1038/sj.onc.1206497

Cited by 14 publications

(11 citation statements)

References 30 publications

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“…Expression of CTAR1-containing constructs, LMP1 and 1-220, was less than LMP1-A5 and 1-220-A5 expression. Lower LMP1 and 1-220 expression is consistent with increased ubiquitination and proteasomal degradation mediated by CTAR1-TRAF binding and consistent with previous expression of these mutants (44,62). In agreement with our previous studies, the levels of p27 protein were decreased to approximately 30% of control p27 levels (17,18,(43)(44)(45).…”

Section: Resultssupporting

confidence: 81%

Transcriptional Downregulation of p27KIP1 through Regulation of E2F Function during LMP1-Mediated Transformation

Everly

Mainou

Raab‐Traub

2009

J Virol

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LMP1 induces the phenotypic transformation of fibroblasts and affects regulators of the cell cycle during this process. LMP1 decreases expression of the cyclin-dependent kinase inhibitor p27 and increases the levels and phosphorylation of cyclin-dependent kinase 2 and the retinoblastoma protein. In the present study, the effects of LMP1 on cell cycle progression and the mechanism of p27 downregulation by LMP1 were determined. Although p27 is frequently regulated at the posttranscriptional level during cell cycle progression and in cancer, LMP1 did not decrease ectopically expressed p27. However, LMP1 did decrease p27 RNA levels and inhibited the activity of p27 promoter reporters. The LMP1-regulated promoter element was mapped to a region containing two E2F sites. Electrophoretic mobility shift assays determined that the regulated cis element bound an inhibitory E2F complex containing E2F4 and p130. These findings indicate that LMP1 decreases p27 transcription through effects on E2F family transcription factors. This property likely contributes to the ability of LMP1 to stimulate cell cycle progression. Epstein-Barr virus (EBV) is a herpesvirus that causes infectious mononucleosis and is associated with a number of malignancies. EBV infection of B cells results in a latent infection and the transformation of B cells in vitro and in vivo.Several proteins are required for EBV-mediated B-cell transformation, including the EBV nuclear antigens (EBNAs), EBNA1, EBNA2, EBNA3A, and EBNA3C, and latent membrane protein 1 (LMP1). LMP1 is expressed in most EBV-associated cancers and is considered the EBV oncogene as it is able to induce fibroblast transformation (34,60,83,84). Expression of LMP1 is frequently detected in nasopharyngeal carcinoma, and LMP1 induces phenotypic changes in epithelial cells and nasopharyngeal carcinoma cell lines, including increased motility, invasion, and migration (12,45,60,66,73).LMP1 is a constitutively active tumor necrosis factor receptor homologue. The transmembrane domain of LMP1 mediates self-association in the absence of ligand, and LMP1 is recruited to cholesterol-rich lipid raft domains in the membrane (3,14,20,28,58,81). LMP1 initiates signaling from two carboxyl-terminal activating regions (CTAR1 and CTAR2) by recruiting tumor necrosis factor receptor-associated factors (TRAFs) and other adaptors (33). LMP1 induces constitutive signaling of nuclear factor B (NF-B), phosphatidylinositol 3-kinase (PI3K), mitogen-activated protein kinase (MAPK), and c-Jun N-terminal kinase (11,16,31,43,44,52,56,61,71).Signaling from CTAR1 is required for EBV B-cell transformation and for LMP1 fibroblast transformation (5,30,32,53,78). CTAR1 uniquely activates several pathways and proteins and induces both canonical and noncanonical NF-B, PI3K, and MAPK signaling. Several growth-stimulating proteins upregulated by CTAR1 have been identified, including the epidermal growth factor receptor, TRAF1, and inhibitors of differentiation 1 (Id1) and Id3 (13,18,51). LMP1 induces the transcription of the epiderma...

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Section: Resultssupporting

confidence: 81%

Transcriptional Downregulation of p27KIP1 through Regulation of E2F Function during LMP1-Mediated Transformation

Everly

Mainou

Raab‐Traub

2009

J Virol

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“…Expression of LMP1 was confirmed using antibodies against the amino-terminal HA tag. The variation in the levels of LMP1 expression has been shown to result from ubiquitination and proteasomal degradation mediated by CTAR1-TRAF binding (44). As reported previously, fulllength LMP1 induced activation of the PI3K-Akt signaling pathway as represented by phosphorylation and inactivation of the Akt target, GSK3␤.…”

Section: Ctar1 Is Necessary For Rodent Fibroblast Transformationmentioning

confidence: 62%

Unique Signaling Properties of CTAR1 in LMP1-Mediated Transformation

Mainou

Everly

Raab‐Traub

2007

J Virol

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The Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) gene is considered the EBV oncogene as it is necessary for EBV-mediated transformation of B lymphocytes and itself transforms rodent fibroblasts. LMP1 activates the NF-B, phosphatidylinositol 3-kinase (PI3K)-Akt, mitogen-activated protein kinase, and Jun N-terminal protein kinase signaling pathways through its two signaling domains, carboxyl-terminal activating regions 1 and 2 (CTAR1 and CTAR2). CTAR1 and CTAR2 induce signal transduction pathways through their direct (CTAR1) or indirect (CTAR2) recruitment of tumor necrosis factor receptor-associated factors (TRAFs). CTAR1 is necessary for LMP1-mediated transformation as well as activation of PI3K signaling and induction of cell cycle markers associated with G 1 /S transition. In this study, activation of PI3K-Akt signaling and deregulation of cell cycle markers were mapped to the TRAF-binding domain within CTAR1 and to the residues between CTAR1 and CTAR2. LMP1 CTAR1 also activated the MEK1/2-extracellular signal-regulated kinase 1/2 signaling pathway, and this activation was necessary for LMP1-induced transformation of Rat-1 fibroblasts. Dominant-negative forms of TRAF2 and TRAF3 inhibited but did not fully block LMP1-mediated transformation. These findings identify a new signaling pathway that is uniquely activated by the TRAF-binding domain of LMP1 and is required for transformation.

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“…Some members of this family of proteins have also been found to ubiquitinate themselves in a RING finger-dependent manner in vivo. Although in some cases (such as the brca1/BARD1 heterodimer (41)), the physiological significance of this activity is not clear, in several others, RING-mediated autoubiquitination has been shown to reduce the stability of the protein in question (42)(43)(44)(45)(46)(47)(48)(49). Here we show that ICP0 provides another example of the principle that E3 ligase autoubiquitination in vivo affects the stability of the E3 ligase itself.…”

Section: Discussionmentioning

confidence: 97%

A RING Finger Ubiquitin Ligase Is Protected from Autocatalyzed Ubiquitination and Degradation by Binding to Ubiquitin-specific Protease USP7

Canning

Boutell

Parkinson

et al. 2004

Journal of Biological Chemistry

128

122

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Herpes simplex virus type 1 immediate-early regulatory protein ICP0 stimulates lytic infection and reactivation from latency, processes that require the ubiquitin E3 ligase activity mediated by the RING finger domain in the N-terminal portion of the protein. ICP0 stimulates the production of polyubiquitin chains by the ubiquitin-conjugating enzymes UbcH5a and UbcH6 in vitro, and in infected and transfected cells it induces the proteasome-dependent degradation of a number of cellular proteins including PML, the major constituent protein of PML nuclear bodies. However, ICP0 binds strongly to the cellular ubiquitin-specific protease USP7, a member of a family of proteins that cleave polyubiquitin chains and/or ubiquitin precursors. The region of ICP0 that is required for its interaction with USP7 has been mapped, and mutations in this domain reduce the functionality of ICP0. These findings pose the question: why does ICP0 include domains that are associated with the potentially antagonistic functions of ubiquitin conjugation and deconjugation? Here we report that although neither protein affected the intrinsic activities of the other in vitro, USP7 protected ICP0 from autoubiquitination in vitro, and their interaction can greatly increase the stability of ICP0 in vivo. These results demonstrate that RING finger-mediated autoubiquitination of ICP0 is biologically relevant and can be regulated by interaction with USP7. This principle may extend to a number of cellular RING finger E3 ubiquitin ligase proteins that have analogous interactions with ubiquitin-specific cleavage enzymes.Herpes simplex virus type 1 (HSV-1) 1 is a common human pathogen that can establish a lifelong quiescent infection in sensory neurons following primary infection of epithelial cells. Environmental stimuli such as stress and sunlight trigger periodic recurrences of lytic infection, causing cold sores and genital lesions. HSV-1 expresses three broad groups of temporally regulated genes during lytic infection, termed immediate-early (IE), early, and late (for reviews, see Refs. 1 and 2). The IE protein ICP0 has an important role in the mechanisms that govern the switch between lytic and latent infection (reviewed in Refs. 3-5). Although not essential for viral replication, ICP0 increases the probability of the virus entering lytic infection, particularly after low multiplicity infection of human fibroblasts, and in its absence, viral genomes are more likely to become repressed and establish a quiescent infection (5-7). ICP0 stimulates the expression of all three classes of viral genes by as yet uncertain mechanisms that correlate with its ability to induce the degradation of a number of cellular proteins (8 -12). ICP0 includes a zinc-binding RING finger domain in its N-terminal portion, and in its C-terminal third lie a nuclear localization signal and motifs required for self-multimerization and efficient localization at specific nuclear substructures known as ND10 or PML nuclear bodies. Consistent with its ability to induce the degradation of...

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Ubiquitination of the Epstein–Barr virus-encoded latent membrane protein 1 depends on the integrity of the TRAF binding site

Cited by 14 publications

References 30 publications

Transcriptional Downregulation of p27KIP1 through Regulation of E2F Function during LMP1-Mediated Transformation

Transcriptional Downregulation of p27KIP1 through Regulation of E2F Function during LMP1-Mediated Transformation

Unique Signaling Properties of CTAR1 in LMP1-Mediated Transformation

A RING Finger Ubiquitin Ligase Is Protected from Autocatalyzed Ubiquitination and Degradation by Binding to Ubiquitin-specific Protease USP7

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