Autophagy is an evolutionarily conserved cellular homeostatic pathway essential for development, immunity, and cell death. Although autophagy modulates MHC antigen presentation, it remains unclear whether autophagy defects impact on CD1d lipid loading and presentation to invariant natural killer T (iNKT) cells and on iNKT cell differentiation in the thymus. Furthermore, it remains unclear whether iNKT and conventional T cells have similar autophagy requirements for differentiation, survival, and/or activation. We report that, in mice with a conditional deletion of the essential autophagy gene Atg7 in the T-cell compartment (CD4 Cre-Atg7), thymic iNKT cell development-unlike conventional T-cell development-is blocked at an early stage and mature iNKT cells are absent in peripheral lymphoid organs. The defect is not due to altered loading of intracellular iNKT cell agonists; rather, it is T-cell-intrinsic, resulting in enhanced susceptibility of iNKT cells to apoptosis. We show that autophagy increases during iNKT cell thymic differentiation and that it developmentally regulates mitochondrial content through mitophagy in the thymus of mice and humans. Autophagy defects result in the intracellular accumulation of mitochondrial superoxide species and subsequent apoptotic cell death. Although autophagy-deficient conventional T cells develop normally, they show impaired peripheral survival, particularly memory CD8 + T cells. Because iNKT cells, unlike conventional T cells, differentiate into memory cells while in the thymus, our results highlight a unique autophagy-dependent metabolic regulation of adaptive and innate T cells, which is required for transition to a quiescent state after population expansion.utophagy is an evolutionarily conserved catabolic process that, by facilitating the breakdown and recycling of damaged organelles and long-lived proteins, is essential to maintaining cellular homeostasis (1). The autophagy pathway is highly regulated during development and also by environmental factors such as nutrient availability/starvation, hypoxia, and reactive oxygen species (ROS). The process is controlled by a number of autophagy-related genes (Atg) and starts with the formation of a double-membraned vesicle (the autophagosome) engulfing the cytoplasmic components to be delivered to the lysosome for degradation. Formation of the autophagosome requires the concerted action of two ubiquitin-like conjugation systems in which Atg12 is covalently linked to Atg5 and Atg8 is conjugated to phosphatidylethanolamine (2, 3). Atg7 is a necessary catalyst in both conjugation systems and is therefore essential for autophagy (3).Due to its important role in cellular homeostasis, dysregulation of autophagy has been implicated in several pathological processes, including neurodegeneration, autoimmunity, cancer, and infection (4). Furthermore, autophagy plays an important role during immune responses by regulating pathogen handling and antigen presentation (5, 6). It is well established that different populations of αβ T lymph...

Ever since it was discovered that central tolerance to self is imposed on developing T cells in the thymus through their interaction with self-peptide major histocompatibility complexes on thymic antigen-presenting cells, immunologists have speculated about the nature of these peptides, particularly in humans. Here, to shed light on the so-far unknown human thymic peptide repertoire, we analyse peptides eluted from isolated thymic dendritic cells, dendritic cell-depleted antigen-presenting cells and whole thymus. Bioinformatic analysis of the 842 identified natural major histocompatibility complex I and II ligands reveals significant crosstalk between major histocompatibility complex-class I and II pathways and differences in source protein representation between individuals as well as different antigen-presenting cells. Furthermore, several autoimmune-and tumour-related peptides, from enolase and vimentin for example, are presented in the healthy thymus. 302 peptides are directly derived from negatively selecting dendritic cells, thus providing the first global view of the peptide matrix in the human thymus that imposes self-tolerance in vivo.

The gene encoding leucine-rich repeat kinase 2 (LRRK2) comprises a major risk factor for Parkinson's disease. Recently, it has emerged that LRRK2 plays important roles in the immune system. LRRK2 is induced by interferon-c (IFNc) in monocytes, but the signaling pathway is not known. Here, we show that IFN-c-mediated induction of LRRK2 was suppressed by pharmacological inhibition and RNA interference of the extracellular signal-regulated kinase 5 (ERK5). This was confirmed by LRRK2 immunostaining, which also revealed that the morphological responses to IFN-c were suppressed by ERK5 inhibitor treatment. Both human acute monocytic leukemia THP-1 cells and human peripheral blood monocytes stimulated the ERK5-LRRK2 pathway after differentiation into macrophages. Thus, LRRK2 is induced via a novel, ERK5-dependent IFN-c signal transduction pathway, pointing to new functions of ERK5 and LRRK2 in human macrophages. Keywords: extracellular signal-regulated kinase, interferon, leucine-rich repeat kinase, macrophages, mitogen-activated protein kinase, signal transduction. M€ uller Str. 27, 72076 T€ ubingen, Germany. E-mail: philipp.kahle@ uni-tuebingen.de Abbreviations used: ERK5, extracellular signal-regulated kinase 5; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GM-CSF, granulocyte-macrophage colony-stimulating factor; HRP, horseradish peroxidase; IFN-c, interferon-c; IN, inhibitor; JAK, Janus kinase; KD, knockdown; LRRK2, leucine-rich repeat kinase 2; MAPK, mitogenactivated kinase; MDM, monocyte-derived macrophage; PD, Parkinson's disease; PMA, phorbol-12-myristate-13-acetate; RNAi, RNA interference; shRNA, small hairpin RNA; STAT1, signal transducer and activator of transcription 1; TBST, Tris-buffered saline + 0.1% Tween 20.

FOXP3-expressing naturally occurring CD4+CD25high T regulatory cells (Treg) are relevant in the control of autoimmunity, and a defect in this cell population has been observed in several human autoimmune diseases. We hypothesized that altered functions of peripheral Treg cells might play a role in the immunopathogenesis of myasthenia gravis, a T cell-dependent autoimmune disease characterized by the presence of pathogenic autoantibodies specific for the nicotinic acetylcholine receptor. We report in this study a significant decrease in the in vitro suppressive function of peripheral Treg cells isolated from myasthenia patients in comparison to those from healthy donors. Interestingly, Treg cells from prednisolone-treated myasthenia gravis patients showed an improved suppressive function compared with untreated patients, suggesting that prednisolone may play a role in the control of the peripheral regulatory network. Indeed, prednisolone treatment prevents LPS-induced maturation of monocyte-derived dendritic cells by hampering the up-regulation of costimulatory molecules and by limiting secretion of IL-12 and IL-23, and enhancing IL-10. In addition, CD4+ T cells cultured in the presence of such tolerogenic dendritic cells are hyporesponsive and can suppress autologous CD4+ T cell proliferation. The results shown in this study indicate that prednisolone treatment promotes an environment that favors immune regulation rather than inflammation.