EBV latency III immortalization program sensitizes B cells to induction of CD95-mediated apoptosis via LMP1: role of NF- B, STAT1, and p53

Abstract

Epstein-Barr virus (EBV) induces IntroductionEpstein-Barr virus (EBV) was first reported to be associated with Burkitt lymphoma (BL). EBV is able to immortalize B cells both in vitro and in vivo. EBV is also the etiologic agent of infectious mononucleosis, a benign viral disease of young adults that regresses spontaneously. In most cases, primary EBV infection is asymptomatic. Epidemiologic and serologic studies showed that EBV is widely distributed in humans around the world and that more than 95% of adults have been infected by EBV in their past. The ability to grow EBV-infected B-cell lines from peripheral blood mononuclear cells (PBMCs) of EBV-seroconverted healthy subjects leads to the conclusion that this virus persists as a clinically silent infection for the lifetime of the infected host. 1 Nevertheless, it is of note that the virus remains continuously active in immunocompetent individuals even though at very low levels. 2 The EBV genome consists of a 180-kb double-stranded DNA that remains stable in the cell nucleus in an episomal form. In vitro immortalization of B cells by EBV is due to the expression of a restricted set of viral genes, the so-called latency III program, coding for latent membrane proteins (LMP1, LMP2a, LMP2b), Epstein-Barr nuclear antigens (EBNA1, 2, 3A, 3B, 3C, LP), and small noncoding RNAs (EBERs), and leads to the establishment of lymphoblastoid cell lines (LCLs). 3 Among these genes, EBNA1 is responsible for episomal maintenance of the EBV genome. 4 EBNA2 alone is responsible for the expression of the entire set of genes of the latency III program. 5 EBNA2 reroutes the Notch pathway by targeting RBP-jkappa. 6 EBNA2 is also the main viral transactivator of the BNLF1 gene coding for LMP1. LMP1 mimics a constitutively activated receptor of the TNF receptor family through aggregation of TRAF/TRADD molecules. 7,8 In vivo, the latency III program is expressed in newly infected B-blasts during primary EBV infection. 9 Most of these newly EBV-infected B-blasts are killed by cytotoxic T cells. However, some EBVinfected B-blasts are considered to follow the normal process of B-cell maturation until the B-cell memory stage. 10 The B-cell memory compartment constitutes the EBV reservoir in humans. EBV is almost silent in its reservoir. [10][11][12] The immune response against EBV is associated with a dramatic oligoclonal increase in EBV-specific CD8 cytotoxic T cells during the initial phase of primary EBV infection. [13][14][15] Cytotoxic T cells may prevent in vitro de novo infection of B cells. 16,17 In Purtilo syndrome, the defect of SH2D1A/SAP gene alters T-cell signaling in response to antigen. 18 The SAP signaling For personal use only. on May 10, 2018. by guest www.bloodjournal.org From pathway plays a key role in the cytolytic activity of T cells against EBV-positive targets, 19 and Purtilo syndrome is characterized by fatal infectious mononucleosis after primo-infection. 18 Emergence of EBV-associated lymphoma in patients with acquired T-cell immunodeficiencies such as HIV infecti...

EBV Can Protect Latently Infected B Cell Lymphomas from Death Receptor-Induced Apoptosis

“…Although the authors dismiss the significance of this association by documenting TRAIL resistance in both EBV Ϫ and EBV ϩ Akata BL clones, their experiments do not address how EBV infection modulates apoptosis in a TRAIL-sensitive cell line such as BJAB. More recently, Tafuku et al (40) (42).…”

Identification of a novel p53-dependent activation pathway of STAT1 by antitumour genotoxic agents

“…Western blotting was performed as described elsewhere, 36 except for detection of phosphotyrosine-701 STAT1. In the latter case, sonicated cellular extracts were loaded in the gel without boiling to preserve the integrity of the phosphate residue on Y701-STAT1.…”

“…Quantitative RT-PCR was performed as described. 36 Total RNA was extracted using the Qiagen kit, following the manufacturer's recommendations. We defined, as reference RNA, a pool of RNAs extracted from different tonsils, lymph nodes and spleen with benign reactive follicular hyperplasia.…”

Regulation of DNA Polymerase β by the LMP1 Oncoprotein of EBV through the Nuclear Factor-κB Pathway

“…Selection of stably transfected cells was performed as previously described (21,23). Forty eight hours after the addition of doxycyclin at 1 μg/mL, NGFR-t inducible cells were purified as described using magnetic beads (22). Control (Ba/F3-neo) and LMP1-variants-transfected Ba/F3 cells (Ba/F3-B95.8/HL1/HL2) were obtained as previously described (20).…”

“…The LMP1 coding sequences from B95.8 LCL and two Hodgkin's lymphoma infiltrated lymph nodes (HL1 and HL2) were amplified and inserted into pCR 3.1-Uni vector (Invitrogen) as previously described (18)(19)(20). The pRT-1 doxycyclin double inducible episomal vector for truncated NGFR (NGFR-t) and a cDNA of interest is described elsewhere (21,22). The cDNAs coding for IκBα S32,36A (constitutively active form of IκBα; ref.…”