Epstein-Barr Virus LMP2A Alters In Vivo and In Vitro Models of B-Cell Anergy, but Not Deletion, in Response to Autoantigen

Abstract

A significant percentage of the population is latently infected with Epstein-Barr virus (EBV). In most individuals, no clinical symptoms are evident. However, in a subset of individuals with compromised immune systems, EBV infection is associated with lymphomas, such as Burkitt's and non-Hodgkin's lymphomas (34, 47). Furthermore, approximately 40% of Hodgkin's lymphoma cases are positive for EBV latent infection, suggesting that there is a causal relationship between EBV infection and the occurrence of Hodgkin's lymphoma (31, 32). Most recently, our laboratory has implicated that an EBVencoded protein, latent membrane protein 2A (LMP2A), alters gene expression in a manner similar to that observed in Hodgkin Reed-Sternberg cells. These data indicate that LMP2A may play a role in the development of Hodgkin's lymphoma (46). In addition, LMP2A expression is readily detected in vivo, suggesting that LMP2A plays a critical role in the EBV life cycle (4,5,14,26). Therefore, understanding the functions of EBV latency proteins may allow for therapeutic interventions to prevent or treat EBV latency and subsequent lymphomas.Studies using human lymphoblastoid cell lines (LCLs) demonstrate that LMP2A induces the constitutive phosphorylation of numerous signal transduction proteins utilized by the B-cell receptor (BCR) (such as Lyn, Syk, and phosphatidylinositol 3-kinase [PI3K]) (19,38,46,49). In these LCLs, LMP2A inhibits the BCR-induced activation of these proteins, suggesting that LMP2A blocks BCR signal transduction in latently infected B cells. However, LCLs are immortalized and actively dividing and express numerous EBV proteins, which is quite different from the EBV latency patterns observed in vivo (3,4,26). Thus, studies of LMP2A function in primary B cells are needed to determine how LMP2A alters normal primary BCR signaling in vivo.In humans, the B cells in which EBV resides are rare (1 in 10 5 to 10 6 B cells) (29, 41, 52), and therefore it is difficult to obtain sufficient numbers of B cells to study how EBV alters their normal function. To address this question, we produced transgenic (Tg) mice that express LMP2A in developing B cells (TgE-Tg) (10, 11). These mice produced immunoglobulin M (IgM)-negative B cells that survive and colonize peripheral organs (11,27). It was proposed that LMP2A acted similarly to a pre-BCR during development since LMP2A decreased the expression of proteins important for rearranging the IgM heavy chain (45) and subsequent heavy-chain rearrangement (11). The survival of these IgM-negative B cells further suggested that LMP2A acted as a BCR mimic, since B cells normally undergo apoptosis from a lack of a tonic BCR-derived survival signal (30, 33). More recently, findings from our laboratory have suggested that LMP2A protects B cells from apoptosis by the RAS/PI3K/AKT pathway (19,46). This model revealed a novel function for LMP2A whereby this latently expressed EBV protein provides a surrogate BCR-like stimulus to promote B-cell development and peripheral B-cell survival. Despite the d...

Syk Tyrosine Kinase Mediates Epstein-Barr Virus Latent Membrane Protein 2A-induced Cell Migration in Epithelial Cells

“…LMP2A can block BCR signaling when expressed in EBV-transformed B cells grown in cell culture (23,47). In contrast, in primary murine B cells LMP2A does not block BCR signaling but alters normal B cell signaling and confers survival and developmental signals to BCR-negative B cells (25,27,48,49). Similarly, Syk might also lead to multiple events in epithelial cells.…”

Epstein-Barr Virus LMP2A Enhances B-Cell Responses In Vivo and In Vitro

“…In these mice, LMP2A is not able to protect B cells from BCR-induced apoptosis in response to autoantigen, suggesting that LMP2A allows BCR signaling to occur. Furthermore, in response to a weaker autoantigen, LMP2A bypassed tolerance induction of B cells by providing additional signals that changed a tolerogenic BCR-induced signal into a functional BCR signal (28). These data suggest that the effect of LMP2A on BCR-derived signals may be positive or negative, depending on the context in which the signals are received.…”

“…Genomic tail DNA was prepared as previously described (4). Primers and the protocol for the PCR were performed as previously described (28).…”

“…In this system, BCR-negative B cells are protected from apoptosis by the LMP2A-mediated activation of the PI3K/Ras pathway (23). More recently, we crossed these LMP2A transgenic mice (TgE) with a strain of mice that expresses a rearranged BCR specific for hen egg lysozyme (HEL-Tg) to generate mice that produce LMP2A-positive B cells with a BCR specific for a known antigen (E/HEL-Tg) (28). In these mice, LMP2A is not able to protect B cells from BCR-induced apoptosis in response to autoantigen, suggesting that LMP2A allows BCR signaling to occur.…”

Murine gammaherpesvirus 68 infection protects lupus-prone mice from the development of autoimmunity

“…This problem has been partly circumvented by generating transgenic mice that express isolated EBV genes which, when used in studies of autoimmunity, have supported a role of EBV in this pathological process (4)(5)(6). Interpretation of these studies, however, has to take into account the absence of the whole viral genome and the infection program, which might have additional effects.…”