A distinct topology of BTN3A IgV and B30.2 domains controlled by juxtamembrane regions favors optimal human γδ T cell phosphoantigen sensing

Abstract Butyrophilin (BTN)–3A and BTN2A1 molecules control the activation of human Vγ9Vδ2 T cells during T cell receptor (TCR)-mediated sensing of phosphoantigens (PAg) derived from microbes and tumors. However, the molecular rules governing PAg sensing remain largely unknown. Here, we establish three mechanistic principles of PAg-mediated γδ T cell activation. First, in humans, following PAg binding to the intracellular BTN3A1-B30.2 domain, Vγ9Vδ2 TCR triggering involves the extracellular V-domain of BTN3A2/BTN3A3. Moreover, the localization of both protein domains on different chains of the BTN3A homo-or heteromers is essential for efficient PAg-mediated activation. Second, the formation of BTN3A homo-or heteromers, which differ in intracellular trafficking and conformation, is controlled by molecular interactions between the juxtamembrane regions of the BTN3A chains. Finally, the ability of PAg not simply to bind BTN3A-B30.2, but to promote its subsequent interaction with the BTN2A1-B30.2 domain, is essential for T-cell activation. Defining these determinants of cooperation and the division of labor in BTN proteins improves our understanding of PAg sensing and elucidates a mode of action that may apply to other BTN family members

Zebrafish Endzone Regulates Neural Crest-Derived Chromatophore Differentiation and Morphology

The development of neural crest-derived pigment cells has been studied extensively as a model for cellular differentiation, disease and environmental adaptation. Neural crest-derived chromatophores in the zebrafish (Danio rerio) consist of three types: melanophores, xanthophores and iridiphores. We have identified the zebrafish mutant endzone (enz), that was isolated in a screen for mutants with neural crest development phenotypes, based on an abnormal melanophore pattern. We have found that although wild-type numbers of chromatophore precursors are generated in the first day of development and migrate normally in enz mutants, the numbers of all three chromatophore cell types that ultimately develop are reduced. Further, differentiated melanophores and xanthophores subsequently lose dendricity, and iridiphores are reduced in size. We demonstrate that enz function is required cell autonomously by melanophores and that the enz locus is located on chromosome 7. In addition, zebrafish enz appears to selectively regulate chromatophore development within the neural crest lineage since all other major derivatives develop normally. Our results suggest that enz is required relatively late in the development of all three embryonic chromatophore types and is normally necessary for terminal differentiation and the maintenance of cell size and morphology. Thus, although developmental regulation of different chromatophore sublineages in zebrafish is in part genetically distinct, enz provides an example of a common regulator of neural crest-derived chromatophore differentiation and morphology

Linking Cancer Metabolism and Innate Immunosurveillance: Modulation of Gamma-Delta T Cell-mediated Tumor Cell Recognition by the Key Metabolic Regulator AMPK

There is accumulating evidence demonstrating that gd T cells comprise an important arm of innate cancer immunosurveillance governed by intracellular accumulation of mevalonate pathway products. In contrast to optimized in vitro conditions, growing tumors in vivo are exposed to nutrient deprivation/hypoxia both activating the key energy metabolism regulator adenosine monophosphate activated protein kinase (AMPK). Upon activation, AMPK increase catabolic ATP-generating processes such as uptake and oxidation of glucose and fatty acids but inhibit ATP-consuming biosynthetic processes such as protein synthesis and cholesterol production (mevalonate pathway). This correlation prompted us to investigate the effects of AMPK activation (mimicked by the synthetic AMP-analogue 5-aminoimidazole-4-carboxamide riboside (=AICAR) or by the anti-diabetic drug metformin on tumor cell recognition by Vg9Vd2 T cells.We found that Vg9Vd2 T cells exhibited significant decreased up-regulation of activation markers in response to AICAR or metformin pre-treated target tumor cell lines (Daudi, RPMI 8226 and the farnesylpyrophphosphate synthase (FPPS)-knockdown cell line Raji AS22) as compared to untreated tumor cells. In addition, AMPK activation in tumor cells reduced the ability of Vg9Vd2 T cells to produce pro-inflammatory cytokines (IFN-g, TNF-a). As determined by phospho-specific flow cytometry analysis, either AICAR or metformin treatment resulted in increased phosphorylation and therefore inactivation of the AMPK target enzyme 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase, subsequently lowering intracellular mevalonate pathway products critical for activation of Vg9Vd2 T cells. Thus, this data demonstrates for the first time an impact of cancer metabolism on immune recognition facilitating tumor escape

Open Access Successful adoptive transfer and in vivo expansion of haploidentical γδ T cells

Background: The primary aim of this pilot study was to determine the feasibility and safety of an adoptive transfer and in vivo expansion of human haploidentical γδ T lymphocytes. Methods: Patients with advanced haematological malignancies who are not eligible for allogeneic transplantation received peripheral blood mononuclear cells from half-matched family donors. For that, a single unstimulated leukapheresis product was incubated with both the anti-CD4 and anti-CD8 antibodies conjugated to paramagnetic particles. The depletion procedure was performed on a fully automated CliniMACS ® device according to the manufacturer’s instructions. On average, patients received 2.17 × 10 6 /kg (range 0.9-3.48) γδ T cells with <1 % CD4or CD8-positive cells remaining in the product. All patients received prior lymphopenia-inducing chemotherapy (fludarabine 20-25 mg/m 2 day-6 until day-2 and cyclophosphamide 30-60 mg/kg day-6 and-5) and were treated with 4 mg zoledronate on day 0 and 1.0x10 6 IU/m 2 IL-2 on day +1 until day +6 for the induction of γδ T cell proliferation in vivo. Results: This resulted in a marked in vivo expansion of donor γδ T cells and, to a lower extent, natural killer cells and double-negative αβ T cells (mean 68-fold, eight-fold, and eight-fold, respectively). Proliferation peaked by around day +8 and donor cells persisted up to 28 days. Although refractory to all prior therapies, three out of four patients achieved a complete remission, which lasted for 8 months in a patient with plasma cell leukaemia. One patient died from an infection 6 weeks after treatment. Conclusion: This pilot study shows that adoptive transfer and in vivo expansion of haploidentical γδ T lymphocytes is feasible and suggests a potential role of these cells in the treatment of haematological diseases

URGI genome annotation system:an integrated system for structural and functional genome annotation

The URGI platform (http://urgi.versailles.inra.fr) develops a genome annotation system dedicated to plants and their pathogens. This Integrated System relies on: (i) pipelines for Transposable Elements annotation (REPET) and gene structural and functional predictions (ii) databases and user-friendly interfaces to browse and query the data (URGI Information System GnpIS, Genome Report System GRS), (iii) A distributed annotation system for curation of gene structure