Latent Epstein-Barr virus infection and the germinal center reaction

Epstein-Barr virus (EBV) is a γ-herpes virus which preferentially infects human B lymphocytes. It is highly adapted to persist in B cells since it encodes for proteins which mimic several cellular proteins playing an important role in B cell biology. Thus, the viral Latent Membrane Proteins (LMP) 1 and 2A are considered to be functional homologues of the CD40 receptor and the B cell receptor, respectively. It has been postulated that EBV uses the normal T cell dependent immune response B cell differentiation pathway (the so-called germinal center reaction) for its infection cycle to gain access to the long living memory B cell compartment. LMP1 and LMP2A are suggested to play an important role during this process, since they are able to provide B cells with survival and proliferation signals, and may help the EBV-infected B cells to evade negative selection during the germinal center reaction. LMP1 and LMP2A are also expressed in germinal center B cell derived EBV-associated malignancies, suggesting a contribution of these viral proteins to tumor development. However, the data concerning the role of the germinal center reaction in the latent EBV infection circle is controversial, remaining it elusive (i) if EBV-infected B cells have to pass the germinal center to establish persistence, and (ii) if LMP1 signaling has any influence on germinal center B cells. In the present work, the interplay of EBV and the germinal center reaction was examined, focusing on the roles of LMP1 and LMP2A in normal B cell biology and lymphomagenesis. LMP1 was shown to mimic a constitutive active CD40 receptor in vitro, but in vivo LMP1 only partially restored the CD40-deficiency in transgenic mice; it even blocked germinal center formation in the presence of CD40. This could be due to differences in the signaling mediated by LMP1 and CD40, or to the constitutive activity of LMP1. To compare CD40 and LMP1 signaling in vivo, we generated a transgenic mouse line which conditionally expresses a CD40-ligand regulated LMP1 protein (CD40/LMP1). We show that LMP1 signaling in B cells perfectly mimics CD40 function in vivo, leading to normal B cell development, B cell activation, and T cell dependent immune responses in CD40 deficient mice. Thus, we conclude that not the LMP1 signaling domain but its constitutive activity interferes with the germinal center reaction. This is in accordance with a previous study of our group, showing that ligand-independent constitutive active CD40 signaling (LMP1/CD40) blocks germinal center initiation like LMP1. However, the influence of a constitutive active CD40 signaling directly on germinal center B cells remained open. In the present study, LMP1/CD40 expression was specifically induced in germinal center B cells. We show that a constitutive active CD40 signaling also interferes with early germinal centers, but leads to lymphoproliferation, which most likely reflects malignant lymphoma. The incidence of 100% suggests a substantial risk of pre-activated B cells to become transformed upon deregulation of CD40 signaling or LMP1 expression. Since constitutive LMP1 and CD40 signals are not compatible with the germinal center B cell differentiation process, and LMP1 expression however has been detected in B cells resembling a germinal center phenotype of healthy EBV-carriers, we questioned whether these are real germinal center B cells. We studied the influence of EBV protein expression on the phenotype of human B cells in vitro, and indeed could show that EBNA2-independent LMP1 and LMP2A expression induces a germinal center cell like phenotype in B cells. Further, the inactivation of EBNA2, the key transactivator of EBV gene expression, does not lead to the original naïve phenotype of a B cell, but induces up-regulation of the memory B cell marker CD27. Beyond, we show that in CD40/LMP1-expressing B cells of the transgenic mice as well as in the human cell lines, LMP1 signaling induces class switch recombination independent from cytokines. This implies a unique feature of the LMP1 signaling domain to initiate differentiation processes beside its ability to activate B cells. Taking these data into account, we suggest that EBV infected B cells do not have to undergo a classical germinal center reaction to enter the memory B cell pool, but that with the help of EBV proteins they are able to induce processes imitating memory B cell differentiation independent from T cells, thus escaping immune surveillance. These EBV-infected quiescent memory-like B cells eventually enter the germinal center reaction upon antigen-dependent activation. There, re-induction of LMP1 might harbor a substantial risk of malignant transformation

Constitutive CD40 signaling in B cells selectively activates the noncanonical NF-κB pathway and promotes lymphomagenesis

CD40, a member of the tumor necrosis factor (TNF) receptor family, plays an essential role in T cell–dependent immune responses. Because CD40 is widely expressed on the surface of tumor cells in various B cell malignancies, deregulated CD40 signaling has been suggested to contribute to lymphomagenesis. In this study, we show that B cell-specific expression of a constitutively active CD40 receptor, in the form of a latent membrane protein 1 (LMP1)/CD40 chimeric protein, promoted an increase in the number of follicular and marginal zone B cells in secondary lymphoid organs in transgenic mice. The B cells displayed an activated phenotype, prolonged survival and increased proliferation, but were significantly impaired in T cell-dependent immune responses. Constitutive CD40 signaling in B cells induced selective and constitutive activation of the noncanonical NF-κB pathway and the mitogen-activated protein kinases Jnk and extracellular signal–regulated kinase. LMP1/CD40-expressing mice older than 12 mo developed B cell lymphomas of mono- or oligoclonal origin at high incidence, thus showing that the interplay of the signaling pathways induced by constitutive CD40 signaling is sufficient to initiate a tumorigenic process, ultimately leading to the development of B cell lymphomas

A source of entangled photons based on a cavity-enhanced and strain-tuned GaAs quantum dot

A quantum-light source that delivers photons with a high brightness and a high degree of entanglement is fundamental for the development of efficient entanglement-based quantum-key distribution systems. Among all possible candidates, epitaxial quantum dots are currently emerging as one of the brightest sources of highly entangled photons. However, the optimization of both brightness and entanglement currently requires different technologies that are difficult to combine in a scalable manner. In this work, we overcome this challenge by developing a novel device consisting of a quantum dot embedded in a circular Bragg resonator, in turn, integrated onto a micromachined piezoelectric actuator. The resonator engineers the light-matter interaction to empower extraction efficiencies up to 0.69(4). Simultaneously, the actuator manipulates strain fields that tune the quantum dot for the generation of entangled photons with fidelities up to 0.96(1). This hybrid technology has the potential to overcome the limitations of the key rates that plague current approaches to entanglement-based quantum key distribution and entanglement-based quantum networks. Introductio

A multipair-free source of entangled photons in the solid state

Unwanted multiphoton emission commonly reduces the degree of entanglement of photons generated by non-classical light sources and, in turn, hampers their exploitation in quantum information science and technology. Quantum emitters have the potential to overcome this hurdle but, so far, the effect of multiphoton emission on the quality of entanglement has never been addressed in detail. Here, we tackle this challenge using photon pairs from a resonantly-driven quantum dot and comparing quantum state tomography and second-order coherence measurements as a function of the excitation power. We observe that the relative (absolute) multiphoton emission probability is as low as $p_m= (5.6 \pm 0.6)10^{-4}$ ($p_2= (1.5 \pm 0.3)10^{-6}$) at the maximum source brightness, values that lead to a negligible effect on the degree of entanglement. In stark contrast with probabilistic sources of entangled photons, we also demonstrate that the multiphoton emission probability and the degree of entanglement remain practically unchanged against the excitation power for multiple Rabi cycles, despite we clearly observe oscillations in the second-order coherence measurements. Our results, explained by a theoretical model that we develop to estimate the actual multiphoton contribution in the two-photon density matrix, highlight that quantum dots can be regarded as a multipair-free source of entangled photons in the solid state

LMP1 signaling can replace CD40 signaling in B cells in vivo and has unique features of inducing class-switch recombination to IgG1.

The Epstein-Barr virus (EBV) protein LMP1 is considered to be a functional homologue of the CD40 receptor. However, in contrast to the latter, LMP1 is a constitutively active signaling molecule. To compare B cell-specific LMP1 and CD40 signaling in an unambiguous manner, we generated transgenic mice conditionally expressing a CD40/LMP1 fusion protein, which retained the LMP1 cytoplasmic tail but has lost the constitutive activity of LMP1 and needs to be activated by the CD40 ligand. We show that LMP1 signaling can completely substitute CD40 signaling in B cells, leading to normal B-cell development, activation, and immune responses including class-switch recombination, germinal center formation, and somatic hypermutation. In addition, the LMP1-signaling domain has a unique property in that it can induce class-switch recombination to IgG1 independent of cytokines. Thus, our data indicate that LMP1 has evolved to imitate T-helper cell function allowing activation, proliferation, and differentiation of EBV-infected B cells independent of T cells

Quantum dot technology for quantum repeaters: from entangled photon generation toward the integration with quantum memories

The realization of a functional quantum repeater is one of the major research goals in long-distance quantum communication. Among the different approaches that are being followed, the one relying on quantum memories interfaced with deterministic quantum emitters is considered as one of the most promising solutions. In this work, we focus on the hardware to implement memory-based quantum-repeater schemes that rely on semiconductor quantum dots (QDs) for the generation of polarization entangled photons. Going through the most relevant figures of merit related to efficiency of the photon source, we select significant developments in fabrication, processing and tuning techniques aimed at combining high degree of entanglement with on-demand pair generation, with a special focus on the progress achieved in the representative case of the GaAs system. We proceed to offer a perspective on integration with quantum memories, both highlighting preliminary works on natural–artificial atomic interfaces and commenting a wide choice of currently available and potentially viable memory solutions in terms of wavelength, bandwidth and noise-requirements. To complete the overview, we also present recent implementations of entanglement-based quantum communication protocols with QDs and highlight the next challenges ahead for the implementation of practical quantum networks

Molecular basis of cytotoxicity of Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) in EBV latency III B cells: LMP1 induces type II ligand-independent autoactivation of CD95/Fas with caspase 8-mediated apoptosis.

International audienceThe Epstein-Barr virus (EBV) oncoprotein latent membrane protein 1 (LMP1) is thought to act as the major transforming protein in various cell types, by rerouting the tumor necrosis factor receptor family signaling pathway. Despite this implication in EBV-associated transformation of cells, LMP1 toxicity is a well-known but poorly studied feature, perhaps because it contradicts its role in transformation. We show that LMP1 physiological levels are very heterogeneous and that the highest levels of LMP1 correlate with Fas overexpression and spontaneous apoptosis in lymphoblastoid cell lines (LCLs). To understand the cytotoxic effect of LMP1 in LCLs, we cloned wild-type LMP1 into a doxycycline double-inducible episomal vector pRT-1, with a truncated version of NGFR as a surrogate marker of inducibility. We found that LMP1 overexpression induced apoptosis in LCL B cells, as shown by annexin V labeling, sub-G(1) peak, and poly(ADP ribose) polymerase cleavage. Knocking down Fas expression by small interfering RNA abolished LMP1-induced apoptosis. The absence of detectable levels of Fas ligand mRNA suggested a ligand-independent activation of Fas. LMP1 induced Fas overexpression with its relocalization in lipid raft microdomains of the membrane. Fas immunoprecipitation detected FADD (Fas-associated death domain protein) and caspase 8, suggesting a Fas-dependent formation of the death-inducing signaling complex. Caspases 8, 9, 3, and 7 were activated by LMP1. Caspase 8 activation was associated with BID cleavage and truncated-BID mitochondrial relocalization, consistent with type II apoptosis. Therefore, our results are in agreement with a model where LMP1-dependent NF-kappaB activation induces Fas overexpression and autoactivation that could overwhelm the antiapoptotic effect of NF-kappaB, revealing an ambivalent function of LMP1 in cell survival and programmed cell death