Peripheral B cells latently infected with Epstein–Barr virus display molecular hallmarks of classical antigen-selected memory B cells

Souza, Tatyana A.; Stollar, B. David; Sullivan, John L.; Luzuriaga, Katherine; Thorley‐Lawson, David A.

doi:10.1073/pnas.0509311102

Cited by 96 publications

(78 citation statements)

References 62 publications

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“…However, a phenotypic and functional analysis of latently infected GC cells in the tonsils showed no obvious impact of LMP1 and LMP2 on the behavior of the cells (46). Furthermore, analyses of expressed immunoglobulin genes in latently infected memory B cells found in the periphery suggested that they had undergone selection by antigen (50,51). This led to the conclusion that the potent signaling effects of LMP1 and LMP2 observed with experimental systems were artifacts, produced by constitutive, unregulated expression, that may be more relevant to EBV-associated diseases.…”

Section: Epstein-barr Virus (Ebv) Is a Human Herpesvirus That Establimentioning

confidence: 99%

Germinal Center B Cells Latently Infected with Epstein-Barr Virus Proliferate Extensively but Do Not Increase in Number

Roughan

Torgbor

Thorley‐Lawson

2010

J Virol

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In this study we show that in long-term persistent infection, Epstein-Barr virus (EBV)-infected cells undergoing a germinal center (GC) reaction in the tonsils are limited to the follicles and proliferate extensively. Despite this, the absolute number of infected cells per GC remains small (average of 3 to 4 cells per germinal center; range, 1 to 9 cells), and only about 38 to 55% (average, 45%) of all GCs carry infected cells. The data fit a model where, on average, cells in the GC divide approximately three times; however, only one progeny cell survives to undergo a further three divisions. Thus, a fraction of cells undergo multiple rounds of division without increasing in numbers; i.e., they die at the same rate that they are dividing. We conclude that EBV-infected cells in the GC undergo the extensive proliferation characteristic of GC cells but that the absolute number is limited either by the immune response or by the availability of an essential survival factor. We suggest that this behavior is a relic of the mechanism by which EBV establishes persistence during acute infection. Lastly, the expression of the viral latent protein LMP1 in GC B cells, unlike in vitro, does not correlate directly with the expression of bcl-2 or bcl-6. This emphasizes our claim that observations made regarding the functions of EBV proteins in cell lines or in transgenic mice should be treated with skepticism unless verified in vivo. Epstein-Barr virus (EBV) is a human herpesvirus that establishes a lifetime persistent infection in Ͼ90% of all adults (reviewed in references 21 and 51). The virus persists in rest-ing, latently infected memory B cells that circulate in the periphery and lymph nodes (5; reviewed in references 53 and 55). In these cells the virus is quiescent, at least at the level of viral protein expression (23). This lack of viral proteins is presumably a major reason why these cells are able to persist in the face of a healthy immune response. The other property for which EBV is well known is its ability to infect resting B cells and drive them to become activated proliferating lymphoblasts through the expression of nine latent proteins and several untranslated RNAs (reviewed in reference 45). The latter include a large number of microRNAs (miRNAs) (7,12,17,44).The current model of Epstein-Barr virus persistence holds that the virus drives resting B cells to become activated lymphoblasts so that they can differentiate through a germinal center (GC) reaction to become resting memory B cells (55-57), where the virus persists. The GC is the structure in the follicles of secondary lymphoid organs where antigen-activated B cells undergo a T-cell-dependent immune response (1, 35, 37). The production and maintenance of GCs are absolutely dependent on the expression of the transcription factor bcl-6 (13, 58) and are initiated by the rapid proliferation of antigenspecific B cells. During this expansion phase, B cells divide approximately every 6 to 12 h (2, 37) so that 3 Ϯ 2 founder cells can create a GC of approximatel...

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Section: Epstein-barr Virus (Ebv) Is a Human Herpesvirus That Establimentioning

confidence: 99%

Germinal Center B Cells Latently Infected with Epstein-Barr Virus Proliferate Extensively but Do Not Increase in Number

Roughan

Torgbor

Thorley‐Lawson

2010

J Virol

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“…Infected cells with low levels of LMP1, for example, would be more likely to evade a host's cytotoxic response against LMP1 than those expressing higher levels. The survivors do rapidly yield daughter populations with the full range of levels of LMP1, allowing them to proliferate and eventually yield in vivo the proliferating memory B cells in which EBV resides latently 64 or the proliferating B cells that can rarely evolve into lymphomas.…”

Section: Po4-eif2αmentioning

confidence: 99%

The LMP1 oncogene of EBV activates PERK and the unfolded protein response to drive its own synthesis

Lee

Sugden

2008

Blood

114

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phorylation of eIF2␣, which upregulates activating transcription factor 4 (ATF4) expression. ATF4, in turn, transactivates LMP1's own promoter. LMP1 activates not only PERK but also inositol requiring kinase 1 (IRE1) and ATF6, 3 pathways of the unfolded protein response (UPR). Increasing expression levels of LMP1 induced a dose-dependent increase in IRE1 activity, as measured by its "splicing" of XBP-1. These infected B cells secrete immunoglobins independent of the levels of LMP1, indicating that only a threshold level of XBP-1 is required for the secretion. These findings indicate that LMP1's activation of the UPR is a normal event in a continuum of LMP1's expression that leads both to stimulatory and inhibitory functions and regulates the physiology of EBV-infected B cells in multiple, unexpected modes. IntroductionOncogenes are dominantly acting regulators of cell proliferation and survival. They can be derived from proto-oncogenes by mutations that alter their enzymatic activity, as in the case of Ha-ras in carcinomas, or their levels of expression, as in the case of Bcl-2 in diffuse, large B-cell lymphomas. 1,2 The latent membrane protein 1 (LMP1) oncogene of Epstein-Barr virus (EBV) stands out among the regulators of cell proliferation because it has opposing activities dependent on its own level of expression. 3 These levels vary naturally in individual cells of clonal populations by more than 100-fold such that in any untreated population, some cells are proliferating while others are not.EBV contributes causally to multiple lymphomas and carcinomas. An important part of its oncogenicity in lymphoid cells lies in its ability to infect, induce, and maintain proliferation in B lymphocytes. This proliferation requires signaling by the LMP1 oncogene. 4-6 LMP1's signaling in B cells is in one way similar to that of the cellular CD40 receptor: both activate nuclear factor-B (NF-B), AP-1, and Stat-1 by associating with molecules such as TRAFs and JAK3. 7-15 LMP1's signaling, however, differs fundamentally from CD40, because LMP1 regulates these signaling pathways without itself being regulated by a ligand, as is CD40. When LMP1 is expressed at intermediate levels, it drives signaling through NF-B, for example, to promote cell proliferation. 6,16 When it is expressed at the high end of physiologic levels, it has been shown to induce the phosphorylation of eukaryotic initiation factor 2␣ (eIF2␣), an inhibition of general protein synthesis and cellular cytostasis. 3,[17][18][19] It is thus essential to elucidate the mechanism by which the levels of LMP1 are regulated to understand its functioning as an oncogene.We examined the path by which LMP1 induces the phosphorylation of eIF2␣ and have found that LMP1 activates the kinase, PERK, which phosphorylates eIF2␣. LMP1's activation of PERK is accompanied by its activation of the inositol requiring kinase 1 (IRE1), and activating transcription factor 6 (ATF6), indicating that LMP1 induces the unfolded protein response (UPR) pathway. LMP1's induction of the UPR is de...

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“…During latency, the virus exists predominantly as a chromatin-associated, multicopy episome in the nuclei of resting B lymphocytes (47,52). Latent infection is associated with several malignancies, including Burkitt's lymphoma, Hodgkin's disease, nasopharyngeal carcinoma, and lymphoproliferative disorders in the immunosuppressed.…”

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Cell Cycle Association of the Retinoblastoma Protein Rb and the Histone Demethylase LSD1 with the Epstein-Barr Virus Latency Promoter Cp

Chau

Deng

Kang

et al. 2008

J Virol

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The Epstein-Barr virus C promoter (Cp) regulates the major multicistronic transcript encoding the EBNA-LP, 1, 2, and 3 genes required for B-cell proliferation during latency. The growth-transforming potential of these viral genes suggests that they must be tightly regulated with the host cell cycle and differentiation process. To better understand Cp regulation, we used DNA affinity purification to identify cellular and viral proteins that bind to Cp in latently infected cells. Several previously unknown factors were identified, including the cell cycle regulatory proteins E2F1 and Rb. E2F1 bound to a specific site in Cp located in the core Cp region 3 of the known EBNA2-responsive RBP-Jk (CSL, CBF1) binding site. The histone H3 K4 demethylase LSD1 (BCC110) was also identified by DNA affinity and was shown to form a stable complex with Rb. Coimmunoprecipitation assays demonstrated that E2F1, Rb, and LSD1 bind to Cp in a cell cycle-dependent manner. Rb and LSD1 binding to Cp increased after the S phase, corresponding to a decrease in histone H3 K4 methylation and Cp transcription. Coimmunoprecipitation and immunofluorescence assays reveal that LSD1 interacts with Rb. Surprisingly, LSD1 did not coimmunoprecipitate with E2F1, suggesting that it associates with Rb independently of E2F1. Depletion of LSD1 by small interfering RNAs inhibited Cp basal transcription levels, and overexpression of LSD1 altered the cell cycle profile in p53-positive (p53 ؉ ), but not p53-negative (p53 ؊ ), HCT cells. These findings indicate that Cp is a cell cycle-regulated promoter that is under the control of Rb and the histone demethylase LSD1 in multiple latency types. Epstein-Barr virus (EBV)is a human gamma-1 herpesvirus that establishes a lifelong latency in over 90% of the world's population (26, 37). During latency, the virus exists predominantly as a chromatin-associated, multicopy episome in the nuclei of resting B lymphocytes (47, 52). Latent infection is associated with several malignancies, including Burkitt's lymphoma, Hodgkin's disease, nasopharyngeal carcinoma, and lymphoproliferative disorders in the immunosuppressed. Different viral-transcription patterns can be observed in each of these malignancies (57). Regulation of the EBV C promoter, or Cp, is important to the biology of EBV because it is the key control point distinguishing latency types and the expression of the viral oncogenes EBNA1, 2, 3A, 3B, 3C, and LP (4). Several cellular transcription factors, including CBF1 (also referred to as CSL or RBP-J,), CBF2 (or Auf1), NF-Y, C/EBP, Sp1, and Egr-1, bind within Cp to regulate its activity (5,17,22,23,34). In addition, the viral protein products of Cp, EBNA-LP, EBNA2, and EBNA3A-C act in conjunction with various cellular factors to autoregulate Cp transcription (22,30,38,59). Deletion mapping of the Cp region showed that sequences Ϫ433 to Ϫ245 upstream of the Cp initiation site (particularly around Ϫ370) are important for the EBNA2/CBF1/CBF2 response, Ϫ119 to Ϫ112 for C/EBP, Ϫ99 to Ϫ91 for Sp1/Egr-1, and Ϫ71 to Ϫ63 fo...

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Peripheral B cells latently infected with Epstein–Barr virus display molecular hallmarks of classical antigen-selected memory B cells

Abstract: Epstein-Barr virus (EBV) establishes a lifelong persistent infection

Cited by 96 publications

References 62 publications

Germinal Center B Cells Latently Infected with Epstein-Barr Virus Proliferate Extensively but Do Not Increase in Number

Germinal Center B Cells Latently Infected with Epstein-Barr Virus Proliferate Extensively but Do Not Increase in Number

The LMP1 oncogene of EBV activates PERK and the unfolded protein response to drive its own synthesis

Cell Cycle Association of the Retinoblastoma Protein Rb and the Histone Demethylase LSD1 with the Epstein-Barr Virus Latency Promoter Cp

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