Goldfarb Y. & Abramson J.
(2024)
Cell Research.
34,
8,
p. 537-538
Autoimmune regulator (AIRE) is a unique transcriptional regulator that induces ectopic expression of thousands of tissue-specific genes in the thymus, a step critical for the establishment of immunological tolerance to self. In their recent publication, Fang et al. provide novel mechanistic insights into AIRE\u2019s modus operandi, by highlighting Z-DNA as a key cis-regulatory element, critical for guiding AIRE to its target genes.
Brabec T., Schwarzer M., Kováčová K., Dobešová M., Schierová D., Březina J., Pacáková I., Šrůtková D., Ben-Nun O., Goldfarb Y., Šplíchalová I., Kolář M., Abramson J., Filipp D. & Dobeš J.
(2024)
Journal of Experimental Medicine.
221,
1,
e20230194.
Intestinal epithelial cells have the capacity to upregulate MHCII molecules in response to certain epithelial-adhesive microbes, such as segmented filamentous bacteria (SFB). However, the mechanism regulating MHCII expression as well as the impact of epithelial MHCII-mediated antigen presentation on T cell responses targeting those microbes remains elusive. Here, we identify the cellular network that regulates MHCII expression on the intestinal epithelium in response to SFB. Since MHCII on the intestinal epithelium is dispensable for SFB-induced Th17 response, we explored other CD4+ T cell-based responses induced by SFB. We found that SFB drive the conversion of cognate CD4+ T cells to granzyme+ CD8α+ intraepithelial lymphocytes. These cells accumulate in small intestinal intraepithelial space in response to SFB. Yet, their accumulation is abrogated by the ablation of MHCII on the intestinal epithelium. Finally, we show that this mechanism is indispensable for the SFB-driven increase in the turnover of epithelial cells in the ileum. This study identifies a previously uncharacterized immune response to SFB, which is dependent on the epithelial MHCII function.
Abramson J., Dobeš J., Lyu M. & Sonnenberg G. F.
(2024)
Nature Reviews Immunology.
24,
p. 64-77
Antigen-presenting cells (APCs) are master regulators of the immune response by directly interacting with T cells to orchestrate distinct functional outcomes. Several types of professional APC exist, including conventional dendritic cells, B cells and macrophages, and numerous other cell types have non-classical roles in antigen presentation, such as thymic epithelial cells, endothelial cells and granulocytes. Accumulating evidence indicates the presence of a new family of APCs marked by the lineage-specifying transcription factor retinoic acid receptor-related orphan receptor-γt (RORγt) and demonstrates that these APCs have key roles in shaping immunity, inflammation and tolerance, particularly in the context of host–microorganism interactions. These RORγt+ APCs include subsets of group 3 innate lymphoid cells, extrathymic autoimmune regulator-expressing cells and, potentially, other emerging populations. Here, we summarize the major findings that led to the discovery of these RORγt+ APCs and their associated functions. We discuss discordance in recent reports and identify gaps in our knowledge in this burgeoning field, which has tremendous potential to advance our understanding of fundamental immune concepts.
Tennenhouse A., Khmelnitsky L., Khalaila R., Yeshaya N., Noronha A., Lindzen M., Makowski E. K., Zaretsky I., Sirkis Y. F., Galon-Wolfenson Y., Tessier P. M., Abramson J., Yarden Y., Fass D. & Fleishman S. J.
(2024)
Nature Biomedical Engineering.
8,
1,
p. 30-44
Conventional methods for humanizing animal-derived antibodies involve grafting their complementarity-determining regions onto homologous human framework regions. However, this process can substantially lower antibody stability and antigen-binding affinity, and requires iterative mutational fine-tuning to recover the original antibody properties. Here we report a computational method for the systematic grafting of animal complementarity-determining regions onto thousands of human frameworks. The method, which we named CUMAb (for computational human antibody design; available at http://CUMAb.weizmann.ac.il), starts from an experimental or model antibody structure and uses Rosetta atomistic simulations to select designs by energy and structural integrity. CUMAb-designed humanized versions of five antibodies exhibited similar affinities to those of the parental animal antibodies, with some designs showing marked improvement in stability. We also show that (1) non-homologous frameworks are often preferred to highest-homology frameworks, and (2) several CUMAb designs that differ by dozens of mutations and that use different human frameworks are functionally equivalent.
Nevo S., Frenkel N., Kadouri N., Gome T., Rosenthal N., Givony T., Avin A., Peligero Cruz C., Kedmi M., Lindzen M., Ben Dor S., Damari G., Porat Z., Haffner-Krausz R., Keren-Shaul H., Yarden Y., Munitz A., Leshkowitz D., Goldfarb Y. & Abramson J.
(2024)
Science Immunology.
9,
91,
eabq6930.
The thymus is a primary lymphoid organ that is essential for the establishment of adaptive immunity through generation of immunocompetent T cells. In response to various stress signals, the thymus undergoes acute but reversible involution. However, the mechanisms governing its recovery are incompletely understood. Here, we used a dexamethasone-induced acute thymic involution mouse model to investigate how thymic hematopoietic cells (excluding T cells) contribute to thymic regeneration. scRNA-seq analysis revealed marked transcriptional and cellular changes in various thymic populations and highlighted thymus-resident innate lymphoid cells type 2 (ILC2) as a key cell type involved in the response to damage. We identified that ILC2 are activated by the alarmins IL-25 and IL-33 produced in response to tissue damage by thymic tuft cells and fibroblasts, respectively. Moreover, using mouse models deficient in either tuft cells and/or IL-33, we found that these alarmins are required for effective thymus regeneration after dexamethasone-induced damage. We also demonstrate that upon their damage-dependent activation, thymic ILC2 produce several effector molecules linked to tissue regeneration, such as amphiregulin and IL-13, which in turn promote thymic epithelial cell differentiation. Collectively, our study elucidates a previously undescribed role for thymic tuft cells and fibroblasts in thymus regeneration through activation of the type 2 immune response.
Gruper Y., Wolff A. S. B., Glanz L., Spoutil F., Marthinussen M. C., Osickova A., Herzig Y., Goldfarb Y., Aranaz-Novaliches G., Dobeš J., Kadouri N., Ben-Nun O., Binyamin A., Lavi B., Givony T., Khalaila R., Gome T., Wald T., Mrazkova B., Sochen C., Besnard M., Ben-Dor S., Feldmesser E., Orlova E. M., Hegedűs C., Lampé I., Papp T., Felszeghy S., Sedlacek R., Davidovich E., Tal N., Shouval D. S., Shamir R., Guillonneau C., Szondy Z., Lundin K. E. A., Osicka R., Prochazka J., Husebye E. S. & Abramson J.
(2023)
Nature.
624,
7992,
p. 653-662
Ameloblasts are specialized epithelial cells in the jaw that have an indispensable role in tooth enamel formation\u2014amelogenesis1. Amelogenesis depends on multiple ameloblast-derived proteins that function as a scaffold for hydroxyapatite crystals. The loss of function of ameloblast-derived proteins results in a group of rare congenital disorders called amelogenesis imperfecta2. Defects in enamel formation are also found in patients with autoimmune polyglandular syndrome type-1 (APS-1), caused by AIRE deficiency3,4, and in patients diagnosed with coeliac disease5\u20137. However, the underlying mechanisms remain unclear. Here we show that the vast majority of patients with APS-1 and coeliac disease develop autoantibodies (mostly of the IgA isotype) against ameloblast-specific proteins, the expression of which is induced by AIRE in the thymus. This in turn results in a breakdown of central tolerance, and subsequent generation of corresponding autoantibodies that interfere with enamel formation. However, in coeliac disease, the generation of such autoantibodies seems to be driven by a breakdown of peripheral tolerance to intestinal antigens that are also expressed in enamel tissue. Both conditions are examples of a previously unidentified type of IgA-dependent autoimmune disorder that we collectively name autoimmune amelogenesis imperfecta.
Oftedal B. E., Berger A. H., Bruserud Ø., Goldfarb Y., Sulen A., Breivik L., Hellesen A., Ben-Dor S., Haffner-Krausz R., Knappskog P. M., Johansson S., Wolff A. S., Bratland E., Abramson J. & Husebye E. S.
(2023)
The Journal of Clinical Investigation.
133,
21,
e169704.
Autoimmune polyendocrine syndrome type 1 (APS-1) is caused by mutations in the autoimmune regulator (AIRE) gene. Most patients present with severe chronic mucocutaneous candidiasis and organ-specific autoimmunity from early childhood, but the clinical picture is highly variable. AIRE is crucial for negative selection of T cells, and scrutiny of different patient mutations has previously highlighted many of its molecular mechanisms. In patients with a milder adult-onset phenotype sharing a mutation in the canonical donor splice site of intron 7 (c.879+1G>A), both the predicted altered splicing pattern with loss of exon 7 (AireEx7\u2013/\u2013) and normal full-length AIRE mRNA were found, indicating leaky rather than abolished mRNA splicing. Analysis of a corresponding mouse model demonstrated that the AireEx7\u2013/\u2013 mutant had dramatically impaired transcriptional capacity of tissue-specific antigens in medullary thymic epithelial cells but still retained some ability to induce gene expression compared with the complete loss-of-function AireC313X\u2013/\u2013 mutant. Our data illustrate an association between AIRE activity and the severity of autoimmune disease, with implications for more common autoimmune diseases associated with AIRE variants, such as primary adrenal insufficiency, pernicious anemia, type 1 diabetes, and rheumatoid arthritis.
Givony T., Leshkowitz D., Del Castillo D., Nevo S., Kadouri N., Dassa B., Gruper Y., Khalaila R., Ben-Nun O., Gome T., Dobeš J., Ben-Dor S., Kedmi M., Keren-Shaul H., Heffner-Krausz R., Porat Z., Golani O., Addadi Y., Brenner O., Lo D. D., Goldfarb Y. & Abramson J.
(2023)
Nature.
622,
7981,
p. 164–172
Development of immunocompetent T cells in the thymus is required for effective defence against all types of pathogens, including viruses, bacteria and fungi. To this end, T cells undergo a very strict educational program in the thymus, during which both non-functional and self-reactive T cell clones are eliminated by means of positive and negative selection 1.Thymic epithelial cells (TECs) have an indispensable role in these processes, and previous studies have shown the notable heterogeneity of these cells 2–7. Here, using multiomic analysis, we provide further insights into the functional and developmental diversity of TECs in mice, and reveal a detailed atlas of the TEC compartment according to cell transcriptional states and chromatin landscapes. Our analysis highlights unconventional TEC subsets that are similar to functionally well-defined parenchymal populations, including endocrine cells, microfold cells and myocytes. By focusing on the endocrine and microfold TEC populations, we show that endocrine TECs require Insm1 for their development and are crucial to maintaining thymus cellularity in a ghrelin-dependent manner; by contrast, microfold TECs require Spib for their development and are essential for the generation of thymic IgA+ plasma cells. Collectively, our study reveals that medullary TECs have the potential to differentiate into various types of molecularly distinct and functionally defined cells, which not only contribute to the induction of central tolerance, but also regulate the homeostasis of other thymus-resident populations.
Kadouri N., Giladi T., Katz S., Hey J., Ben-Dor S., Damari G., Dassa B., Dobes J., Dieter W., Bähr M., Paulsen M., Haffner-Krausz R., Mall M. A., Plass C., Kafka Y. & Abramson J.
(2022)
Science Immunology.
7,
74,
eabn8144.
FOXN1 is a transcription factor critical for the development of both thymic epithelial cell (TEC) and hair follicle cell (HFC) compartments. However, mechanisms controlling its expression remain poorly understood. To address this question, we performed thorough analyses of the evolutionary conservation and chromatin status of the Foxn1 locus in different tissues and states and identified several putative cis-regulatory regions unique to TECs versus HFCs. Furthermore, experiments using genetically modified mice with specific deletions in the Foxn1 locus and additional bioinformatic analyses helped us identify key regions and transcription factors involved in either positive or negative regulation of Foxn1 in both TECs and HFCs. Specifically, we identified SIX1 and FOXN1 itself as key factors inducing Foxn1 expression in embryonic and neonatal TECs. Together, our data provide important mechanistic insights into the transcriptional regulation of the Foxn1 gene in TEC versus HFC and highlight the role of FOXN1 in its autoregulation.
Dobeš J., Ben-Nun O., Binyamin A., Stoler-Barak L., Oftedal B. E., Goldfarb Y., Kadouri N., Gruper Y., Givony T., Zalayat I., Kováčová K., Böhmová H., Valter E., Shulman Z., Filipp D., Husebye E. S. & Abramson J.
(2022)
Nature Immunology.
23,
7,
p. 1098-1108
Patients with loss of function in the gene encoding the master regulator of central tolerance AIRE suffer from a devastating disorder called autoimmune polyendocrine syndrome type 1 (APS-1), characterized by a spectrum of autoimmune diseases and severe mucocutaneous candidiasis. Although the key mechanisms underlying the development of autoimmunity in patients with APS-1 are well established, the underlying cause of the increased susceptibility to Candida albicans infection remains less understood. Here, we show that Aire+MHCII+ type 3 innate lymphoid cells (ILC3s) could sense, internalize and present C. albicans and had a critical role in the induction of Candida-specific T helper 17 (TH17) cell clones. Extrathymic Rorc-Cre-mediated deletion of Aire resulted in impaired generation of Candida-specific TH17 cells and subsequent overgrowth of C. albicans in the mucosal tissues. Collectively, our observations identify a previously unrecognized regulatory mechanism for effective defense responses against fungal infections.
Heinlein M., Gandolfo L. C., Zhao K., Teh C. E., Nguyen N., Baell J. B., Goldfarb Y., Abramson J., Wichmann J., Voss A. K., Strasser A., Smyth G. K., Thomas T. & Gray D. H. D.
(2022)
Science Immunology.
7,
67,
eabb6032.
The autoimmune regulator (AIRE) induces the transcription of thousands of peripheral tissue genes (PTGs) in thymic epithelial cells (TECs) to mediate immunological tolerance. The chromatin state required for optimal AIRE function in TECs and how this state is induced remains unclear. We tested the role of the histone acetyltransferase, KAT7 (also known as HBO1 or MYST2), which is essential for acetylation of histone 3 lysine 14, in TEC differentiation, AIRE-mediated PTG expression, and thymic tolerance. We find that KAT7 is required for optimal expansion of medullary TEC and has a major role in the expression of AIRE-dependent PTGs, associated with enhanced chromatin accessibility at these gene loci in TECs. Mice with TEC-specific
deletion develop organ-specific autoimmunity with features resembling those observed in
-deficient mice. These findings highlight critical roles for KAT7-mediated acetylation in promoting a chromatin state at PTG loci that enables AIRE function and the establishment of immunological tolerance.
Goldfarb Y., Givony T., Dobeš J., Kadouri N., Zalayat I., Peligero-Cruz C., Damari G., Dassa B., Ben-Dor S., Gruper Y., Oftedal B. E., Bratland E., Erichsen M. M., Berger A., Avin A., Nevo S., Haljasorg U., Kuperman Y., Ulman A., Porat Z., Haffner-Krausz R., Atasoy U., Leshkowitz D., Husebye E. S. & Abramson J.
(2021)
Journal of Experimental Medicine.
218,
11,
e20201076.
The autoimmune regulator (AIRE) is essential for the establishment of central tolerance and prevention of autoimmunity. Interestingly, different AIRE mutations cause autoimmunity in either recessive or dominant-negative manners. Using engineered mouse models, we establish that some monoallelic mutants, including C311Y and C446G, cause breakdown of central tolerance. By using RNAseq, ATACseq, ChIPseq, and protein analyses, we dissect the underlying mechanisms for their dominancy. Specifically, we show that recessive mutations result in a lack of AIRE protein expression, while the dominant mutations in both PHD domains augment the expression of dysfunctional AIRE with altered capacity to bind chromatin and induce gene expression. Finally, we demonstrate that enhanced AIRE expression is partially due to increased chromatin accessibility of the AIRE proximal enhancer, which serves as a docking site for AIRE binding. Therefore, our data not only elucidate why some AIRE mutations are recessive while others dominant, but also identify an autoregulatory mechanism by which AIRE negatively modulates its own expression.
Peligero-Cruz C., Givony T., Sebe-Pedros A., Dobes J., Kadouri N., Nevo S., Roncato F., Alon R., Goldfarb Y. & Abramson J.
(2020)
eLife.
9,
e58213.
Foxp3+ regulatory T cells (Tregs) are potent suppressor cells, essential for the maintenance of immune homeostasis. Most Tregs develop in the thymus and are then released into the immune periphery. However, some Tregs populate the thymus and constitute a major subset of yet poorly understood cells. Here we describe a subset of thymus recirculating IL18R+ Tregs with molecular characteristics highly reminiscent of tissue-resident effector Tregs. Moreover, we show that IL18R+ Tregs are endowed with higher capacity to populate the thymus than their IL18R– or IL18R–/– counterparts, highlighting the key role of IL18R in this process. Finally, we demonstrate that IL18 signaling is critical for the induction of the key thymus-homing chemokine receptor – CCR6 on Tregs. Collectively, this study provides a detailed characterization of the mature Treg subsets in the mouse thymus and identifies a key role of IL18 signaling in controlling the CCR6-CCL20-dependent migration of Tregs into the thymus.
Delacher M., Barra M. M., Herzig Y., Eichelbaum K., Rafiee M., Richards D. M., Traeger U., Hofer A., Kazakov A., Braband K. L., Gonzalez M., Woehrl L., Schambeck K., Imbusch C. D., Abramson J., Krijgsveld J. & Feuerer M.
(2020)
iScience.
23,
5,
101127.
Regulatory T cells are important regulators of the immune system and have versatile functions for the homeostasis and repair of tissues. They express the forkhead box transcription factor Foxp3 as a lineage-defining protein. Negative regulators of Foxp3 expression are not well understood. Here, we generated double-stranded DNA probes complementary to the Foxp3 promoter sequence and performed a pull-down with nuclear protein in vitro, followed by elution of bound proteins and quantitative mass spectrometry. Of the Foxp3-promoter-binding transcription factors identified with this approach, one was T cell factor 1 (TCF1). Using viral over-expression, we identified TCF1 as a repressor of Foxp3 expression. In TCF1-deficient animals, increased levels of Foxp3(intermediate)CD25(negative) T cells were identified. CRISPR-Cas9 knockout studies in primary human and mouse conventional CD4 T (T-conv) cells revealed that TCF1 protects T-conv cells from inadvertent Foxp3 expression. Our data implicate a role of TCF1 in suppressing Foxp3 expression in activated T cells.
Recent studies using single-cell genomic technologies and in vivo fate mapping have shown that thymic epithelial cells are far more heterogeneous than previously thought, comprising multiple subpopulations with distinct molecular and functional characteristics.The generation of a functional T cell repertoire in the thymus is mainly orchestrated by thymic epithelial cells (TECs), which provide developing T cells with cues for their navigation, proliferation, differentiation and survival. The TEC compartment has been segregated historically into two major populations of medullary TECs and cortical TECs, which differ in their anatomical localization, molecular characteristics and functional roles. However, recent studies have shown that TECs are highly heterogeneous and comprise multiple subpopulations with distinct molecular and functional characteristics, including tuft cell-like or corneocyte-like phenotypes. Here, we review the most recent advances in our understanding of TEC heterogeneity from a molecular, functional and developmental perspective. In particular, we highlight the key insights that were recently provided by single-cell genomic technologies and in vivo fate mapping and discuss them in the context of previously published data.
Tuft cells are epithelial chemosensory cells with unique morphological and molecular characteristics, the most noticeable of which is a tuft of long and thick microvilli on their apical side, as well as expression of a very distinct set of genes, including genes encoding various members of the taste transduction machinery and pro-inflammatory cyclooxygenases. Initially discovered in rat trachea, tuft cells were gradually identified in various mucosal tissues, and later also in non-mucosal tissues, most recent of which is the thymus.Although tuft cells were discovered more than 60 years ago, their functions in the various tissues remained a mystery until recent years. Today, tuft cells are thought to function as sensors of various types of chemical signals, to which they respond by secretion of diverse biological mediators such as IL25 or acetylcholine. Intestinal tuft cells were also shown to mediate type 2 immunity against parasites.Here, we review the current knowledge on tuft cell characteristics, development and heterogeneity, discuss their potential functions and explore the possible implications and significance of their discovery in the thymus.
Delacher M., Schmidl C., Herzig Y., Breloer M., Hartmann W., Brunk F., Kaegebein D., Traeger U., Hofer A., Bittner S., Weichenhan D., Imbusch C. D., Hotz-Wagenblatt A., Hielscher T., Breiling A., Federico G., Groene H., Schmid R. M., Rehli M., Abramson J. & Feuerer M.
(2019)
Nature Communications.
10,
1621.
The transcriptional regulator Rbpj is involved in T-helper (T-H) subset polarization, but its function in T-reg cells remains unclear. Here we show that T-reg-specific Rbpj deletion leads to splenomegaly and lymphadenopathy despite increased numbers of T-reg cells with a polyclonal TCR repertoire. A specific defect of Rbpj-deficient T-reg cells in controlling T(H)2 polarization and B cell responses is observed, leading to the spontaneous formation of germinal centers and a T(H)2-associated immunoglobulin class switch. The observed phenotype is environment-dependent and can be induced by infection with parasitic nematodes. Rbpj-deficient T-reg cells adopt open chromatin landscapes and gene expression profiles reminiscent of tissue-derived T(H)2-polarized T-reg cells, with a prevailing signature of the transcription factor Gata-3. Taken together, our study suggests that T-reg cells require Rbpj to specifically restrain T(H)2 responses, including their own excessive T(H)2-like differentiation potential.
Goldfarb Y., Peligero-Cruz C. & Abramson J.
(2019)
The Autoimmune Diseases
.
6 ed.
p. 65-90
T lymphocytes are essential guardians against a plethora of pathogens and are thus characterized by an unprecedented diversity for antigen recognition. Their repertoire diversity is achieved by an elaborate, semirandom rearrangement of the genes encoding their respective antigen-specific receptors which inevitably generates many T cells that recognize not only foreign and potentially harmful antigens, but also the body’s own components. Therefore to deal with this problem and to prevent autoimmune destruction, the immune system has evolved a comprehensive network of several complementary mechanisms that are guided by four major principles to ensure tolerance to the body’s own antigens: (1) complete physical elimination of the self-reactive T-cell clone from the repertoire (clonal deletion), (2) conversion of the self-reactive clone into a tolerogenic and/or harmless T-cell subtype (anergy, phenotype skewing), (3) prevention of T-cell encounter with its specific self-antigen in the immune periphery (ignorance, antigen sequestering), and (4) prevention of clonal expansion or reactivation upon self-antigen (re)encounter (immunosuppression, T-cell intrinsic inhibitory mechanisms). In this chapter, we shall try to provide a fresh look at these key concepts and mechanisms underlying the establishment, maintenance, and breakdown of T-cell tolerance to the body’s own components.
Bornstein C., Nevo S., Giladi A., Kadouri N., Pouzolles M., Gerbe F., David E., Machado A., Chuprin A., Toth B., Goldberg O., Itzkovitz S., Taylor N., Jay P., Zimmermann V. S., Abramson J. & Amit I.
(2018)
Nature.
559,
7715,
p. 622-626
T cell development and selection are coordinated in the thymus by a specialized niche of diverse stromal populations(1-3). Although much progress has been made over the years in identifying the functions of the different cell types of the thymic stromal compartment, there is no comprehensive characterization of their diversity and heterogeneity. Here we combined massively parallel single-cell RNA-sequencing(4,5), spatial mapping, chromatin profiling and gene targeting to characterize de novo the entire stromal compartment of the mouse thymus. We identified dozens of cell states, with thymic epithelial cells (TECs) showing the highest degree of heterogeneity. Our analysis highlights four major medullary TEC (mTEC I-IV) populations, with distinct molecular functions, epigenetic landscapes and lineage regulators. Specifically, mTEC IV constitutes a new and highly divergent TEC lineage with molecular characteristics of the gut chemosensory epithelial tuft cells. Mice deficient in Pou2f3, a master regulator of tuft cells, have complete and specific depletion of mTEC IV cells, which results in increased levels of thymus-resident type-2 innate lymphoid cells. Overall, our study provides a comprehensive characterization of the thymic stroma and identifies a new tuft-like TEC population, which is critical for shaping the immune niche in the thymus.
Avin A., Levy M., Porat Z. & Abramson J.
(2017)
Nature Communications.
8,
1,
1524.
In spite of recent advances in proteomics, quantitative analyses of protein-protein interactions (PPIs) or post-translational modifications (PTMs) in rare cell populations remain challenging. This is in particular true for analyses of rare immune and/or stem cell populations that are directly isolated from humans or animal models, and which are often characterized by multiple surface markers. To overcome these limitations, here we have developed proximity ligation imaging cytometry (PLIC), a protocol for proteomic analysis of rare cells. Specifically, by employing PLIC on medullary thymic epithelial cells (mTECs), which serve as a paradigm for a rare immune population, we demonstrate that PLIC overcomes the inherent limitations of conventional proteomic approaches and enables a high-resolution detection and quantification of PPIs and PTMs at a single cell level.
Sheridan J. M., Keown A., Policheni A., Roesley S. N., Rivlin N., Kadouri N., Ritchie M. E., Jain R., Abramson J., Heng T. S. & Gray D. H.
(2017)
Cell Reports.
21,
4,
p. 934-942
Evidence suggests that a stem-cell-driven differentiation hierarchy maintains the dynamic thymic epithelial cell (TEC) network that governs T lymphocyte development. The identification of TEC stem/progenitor cells has been a major focus in the field, and several candidates with contrasting phenotypes have been described. We sought to determine the provenance and function of the only population reported to exhibit TEC stem cell properties in the adult, a Foxn1− EpCAM− cell that generates so-called thymospheres. We provide evidence that the thymosphere-forming cell (TSFC) is not a TEC stem cell but can incorporate bystander TECs into thymospheres, providing an explanation for the epithelial activity ascribed to these structures. TSFCs were found to share a phenotype, transcriptional profile, and developmental origin with thymic fibroblasts and can generate adipocytes. In summary, this study redefines the nature of bipotent TEC stem/progenitor cells in the adult thymus and highlights a potentially important mesenchymal progenitor population. The phenotype of putative bipotent epithelial stem/progenitor cells in the adult thymus has been controversial. Sheridan et al. find that a prominent candidate, the FoxN1−EpCAM− sphere-forming cell, harbors no epithelial stem/progenitor capacity and present evidence that they are a unique intrathymic mesenchymal stromal/stem cell population with adipocyte differentiation capacity.
Abramson J. & Anderson G.
(2017)
Annual Review of Immunology.
35,
p. 85-118
Intrathymic T cell development is a complex process that depends upon continuous guidance from thymus stromal cell microenvironments. The thymic epithelium within the thymic stroma comprises highly specialized cells with a high degree of anatomic, phenotypic, and functional heterogeneity. These properties are collectively required to bias thymocyte development toward production of self-tolerant and functionally competent T cells. The importance of thymic epithelial cells (TECs) is evidenced by clear links between their dysfunction and multiple diseases where autoimmunity and immunodeficiency are major components. Consequently, TECs are an attractive target for cell therapies to restore effective immune system function. The pathways and molecular regulators that control TEC development are becoming clearer, as are their influences on particular stages of T cell development. Here, we review both historical and the most recent advances in our understanding of the cellular and molecular mechanisms controlling TEC development, function, dysfunction, and regeneration.
Herzig Y., Nevo S., Bornstein C., Brezis M. R., Ben-Hur S., Shkedy A., Eisenberg-Bord M., Levi B., Delacher M., Goldfarb Y., David E., Weinberger L., Viukov S., Ben-Dor S., Giraud M., Hanna J. H., Breiling A., Lyko F., Amit I., Feuerer M. & Abramson J.
(2017)
Nature Immunology.
18,
2,
p. 161-172
Aire is a transcriptional regulator that induces promiscuous expression of thousands of genes encoding tissue-restricted antigens (TRAs) in medullary thymic epithelial cells (mTECs). While the target genes of Aire are well characterized, the transcriptional programs that regulate its own expression have remained elusive. Here we comprehensively analyzed both cis-Acting and trans-Acting regulatory mechanisms and found that the Aire locus was insulated by the global chromatin organizer CTCF and was hypermethylated in cells and tissues that did not express Aire. In mTECs, however, Aire expression was facilitated by concurrent eviction of CTCF, specific demethylation of exon 2 and the proximal promoter, and the coordinated action of several transcription activators, including Irf4, Irf8, Tbx21, Tcf7 and Ctcfl, which acted on mTEC-specific accessible regions in the Aire locus.
Danan-Gotthold M., Guyon C., Giraud M., Levanon E. Y. & Abramson J.
(2016)
GENOME BIOLOGY.
17,
1,
219.
Background: In order to become functionally competent but harmless mediators of the immune system, T cells undergo a strict educational program in the thymus, where they learn to discriminate between self and non-self. This educational program is, to a large extent, mediated by medullary thymic epithelial cells that have a unique capacity to express, and subsequently present, a large fraction of body antigens. While the scope of promiscuously expressed genes by medullary thymic epithelial cells is well-established, relatively little is known about the expression of variants that are generated by co-transcriptional and post-transcriptional processes. Results: Our study reveals that in comparison to other cell types, medullary thymic epithelial cells display significantly higher levels of alternative splicing, as well as A-to-I and C-to-U RNA editing, which thereby further expand the diversity of their self-antigen repertoire. Interestingly, Aire, the key mediator of promiscuous gene expression in these cells, plays a limited role in the regulation of these transcriptional processes. Conclusions: Our results highlight RNA processing as another layer by which the immune system assures a comprehensive self-representation in the thymus which is required for the establishment of self-tolerance and prevention of autoimmunity.
Abramson J. & Husebye E. S.
(2016)
Immunological Reviews.
271,
1,
p. 127-140
The establishment of central tolerance in the thymus is critical for avoiding deleterious autoimmune diseases. Autoimmune regulator (AIRE), the causative gene in autoimmune polyendocrine syndrome type-1 (APS-1), is crucial for the establishment of self-tolerance in the thymus by promoting promiscuous expression of a wide array of tissue-restricted self-antigens. This step is critical for elimination of high-affinity self-reactive T cells from the immunological repertoire, and for the induction of a specific subset of Foxp3+ T-regulatory (Treg) cells. In this review, we discuss the most recent advances in our understanding of how AIRE operates on molecular and cellular levels, as well as of how its loss of function results in breakdown of self-tolerance mechanisms characterized by a broad and heterogeneous repertoire of autoimmune phenotypes.
Goldfarb Y., Kadouri N., Levi B., Sela A., Herzig Y., Cohen R. N., Hollenberg A. N. & Abramson J.
(2016)
Cell Reports.
15,
3,
p. 651-665
The thymus provides a unique microenvironment enabling development and selection of T lymphocytes. Medullary thymic epithelial cells (mTECs) play a pivotal role in this process by facilitating negative selection of self-reactive thymocytes and the generation of Foxp3+ regulatory T cells. Although studies have highlighted the non-canonical nuclear factor κB (NF-κB) pathway as the key regulator of mTEC development, comprehensive understanding of the molecular pathways regulating this process still remains incomplete. Here, we demonstrate that the development of functionally competent mTECs is regulated by the histone deacetylase 3 (Hdac3). Although histone deacetylases are global transcriptional regulators, this effect is highly specific only to Hdac3, as neither Hdac1 nor Hdac2 inactivation caused mTEC ablation. Interestingly, Hdac3 induces an mTEC-specific transcriptional program independently of the previously recognized RANK-NFκB signaling pathway. Thus, our findings uncover yet another layer of complexity of TEC lineage divergence and highlight Hdac3 as a major and specific molecular switch crucial for mTEC differentiation.
Abramson J. & Goldfarb Y.
(2016)
European Journal of Immunology.
46,
1,
p. 22-33
Autoimmune regulator (AIRE) is a unique transcriptional regulator that induces promiscuous expression of thousands of tissue-restricted antigens (TRAs) in medullary thymic epithelial cells (mTECs), a step critical for the induction of immunological self-tolerance. The past 15 years have seen dramatic progress in our understanding of how AIRE induces immunological self-tolerance on a molecular level. This major advancement can be greatly attributed to the identification of a large variety of proteins that physically associate with AIRE, supporting and regulating its transcription-transactivation capacity. These diverse molecular partnerships have been shown to play roles in shuttling AIRE to the nucleus, securing AIRE's interaction with nuclear matrix and chromatin, releasing RNA polymerase-II from its stalled state and potentiating AIRE-mediated gene expression, among others. In this review we discuss the relationship of AIRE with its vast and rather diverse repertoire of partners and highlight how such "promiscuous partnerships" contribute to the phenomenon of "promiscuous gene expression" in the thymus.
Satoh R., Kakugawa K., Yasuda T., Yoshida H., Sibilia M., Katsura Y., Levi B., Abramson J., Koseki Y., Koseki H., van Ewijk E. W., Hollander G. A. & Kawamoto H.
(2016)
PLoS Genetics.
12,
1,
e1005776.
Thymic medullary regions are formed in neonatal mice as islet-like structures, which increase in size over time and eventually fuse a few weeks after birth into a continuous structure. The development of medullary thymic epithelial cells (TEC) is dependent on NF-κB associated signaling though other signaling pathways may contribute. Here, we demonstrate that Stat3-mediated signals determine medullary TEC cellularity, architectural organization and hence the size of the medulla. Deleting Stat3 expression selectively in thymic epithelia precludes the postnatal enlargement of the medulla retaining a neonatal architecture of small separate medullary islets. In contrast, loss of Stat3 expression in cortical TEC neither affects the cellularity or organization of the epithelia. Activation of Stat3 is mainly positioned downstream of EGF-R as its ablation in TEC phenocopies the loss of Stat3 expression in these cells. These results indicate that Stat3 meditated signal via EGF-R is required for the postnatal development of thymic medullary regions.
Chuprin A., Sagi I., Avin A., Goldfarb Y., Herzig Y., Levi B., Jacob A., Sela A., Katz S., Grossman M., Guyon C., Rathaus M., Cohen H. Y., Sagi I., Giraud M., McBurney M. W., Husebye E. S. & Abramson J.
(2015)
Nature Immunology.
16,
7,
p. 737-745
Aire is a transcriptional regulator that induces the promiscuous expression of thousands of tissue-restricted antigens (TRAs) in medullary thymic epithelial cells (mTECs), a step critical for the induction of immunological self-tolerance. Studies have offered molecular insights into how Aire operates, but more comprehensive understanding of this process still remains elusive. Here we found abundant expression of the protein deacetylase Sirtuin-1 (Sirt1) in mature Aire + mTECs, wherein it was required for the expression of Aire-dependent TRA-encoding genes and the subsequent induction of immunological self-tolerance. Our study elucidates a previously unknown molecular mechanism for Aire-mediated transcriptional regulation and identifies a unique function for Sirt1 in preventing organ-specific autoimmunity.
Oftedal B., Hellesen A., Erichsen M., Bratland E., Vardi A., Perheentupa J., Kemp E., Fiskerstrand T., Viken M., Weetman A., Fleishman S., Banka S., Newman W., Sewell W., Sozaeva L., Zayats T., Haugarvoll K., Orlova E., Haavik J., Johansson S., Knappskog P., Lovas K., Wolff A., Abramson J. & Husebye E.
(2015)
Immunity.
42,
6,
p. 1185-1196
The autoimmune regulator (AIRE) gene is crucial forestablishing central immunological tolerance and preventing autoimmunity. Mutations in AIRE cause a rare autosomal-recessive disease, autoimmune polyendocrine syndrome type 1 (APS-1), distinguished by multi-organ autoimmunity. We have identified multiple cases and families with mono-allelic mutations in the first plant homeodomain (PHD1) zinc finger of AIRE that followed dominant inheritance, typically characterized by later onset, milder phenotypes, and reduced penetrance compared to classical APS-1. These missense PHD1 mutations suppressed gene expression driven by wild-type AIRE in a dominant-negative manner, unlike CARD or truncated AIRE mutants that lacked such dominant capacity. Exome array analysis revealed that the PHD1 dominant mutants were found with relatively high frequency (>0.0008) in mixed populations. Our results provide insight into the molecular action of AIRE and demonstrate that disease-causing mutations in the AIRE locus are more common than previously appreciated and cause more variable autoimmune phenotypes.
Richards D. M., Delacher M., Goldfarb Y., Kägebein D., Hofer A. C., Abramson J. & Feuerer M.
(2015)
Regulatory T Cells in Health and Disease, 2015
.
Liston A.(eds.).
p. 175-205
Regulatory T cells (Tregs) are crucial mediators of self-tolerance in the periphery. They differentiate in the thymus, where interactions with thymus-resident antigen-presenting cells, an instructive cytokine milieu, and stimulation of the T cell receptor lead to the selection into the Treg lineage and the induction of Foxp3 gene expression. Once mature, Treg cells leave the thymus and migrate into either the secondary lymphoid tissues, e.g., lymph nodes and spleen, or peripheral nonlymphoid tissues. There is growing evidence that Treg cells go beyond the classical modulation of immune responses and also play important functional roles in nonlymphoid peripheral tissues. In this review, we summarize recent findings about the thymic Treg lineage differentiation as well as the further specialization of Treg cells in the secondary lymphoid and in the peripheral nonlymphoid organs.