Antigen processing and presentation in the thymus: implications for T cell repertoire selection

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ScienceDirect Antigen processing and presentation in the thymus: implications for T cell repertoire selection Kenta Kondo1, Kensuke Takada1,2 and Yousuke Takahama1 The processing and presentation of major histocompatibility complex (MHC)-associated antigens depend on the intracellular digestion of self- and nonself-proteins, the loading of digested peptides onto MHC molecules, and the traffic of peptide–MHC complexes to plasma membrane surface for display to interacting T cells. Recent studies have revealed unique machineries for antigen processing and presentation in thymic antigen-presenting cells that display self-antigens to developing thymocytes for the formation of functionally competent yet self-tolerant T cell repertoire. Here, we briefly summarize those machineries, focusing on the biology of cortical and medullary thymic epithelial cells. Addresses 1 Division of Experimental Immunology, Institute of Advanced Medical Sciences, University of Tokushima, 3-18-15 Kuramoto, Tokushima 7708503, Japan 2 Laboratory of Molecular Medicine, Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, N18W9 Kita-ku, Sapporo 060-0818, Japan Corresponding author: Takahama, Yousuke (takahama@genome. tokushima-u.ac.jp)

Current Opinion in Immunology 2017, 46:53–57 This review comes from a themed issue on Antigen processing Edited by Peter Cresswell and Paul A Roche

http://dx.doi.org/10.1016/j.coi.2017.03.014 0952-7915/ã 2017 Elsevier Ltd. All rights reserved.

Introduction The thymus is an organ that produces a pool of T cells that are capable of responding to diverse foreign antigens and that are tolerant to self-antigens. The thymus attracts hematopoietic stem cell derived T-lymphoid progenitors and induces their differentiation into T cell antigen receptor (TCR)-expressing CD4+CD8+ double-positive (DP) thymocytes in the microenvironment of the thymic cortex [1,2]. The antigen recognition specificity of TCRs expressed by individual DP thymocytes is determined cell-by-cell through the V(D)J rearrangement of the genome in the nucleus, so that the de novo repertoire of TCRs expressed by newly generated DP thymocytes contains a wide variety of TCR specificities, including www.sciencedirect.com

TCRs that are unable to interact with self-major histocompatibility complex (MHC)-associated foreign antigens, that is, useless TCR specificities; or TCRs that are highly reactive with self-MHC-associated self-antigens, that is, harmful TCR specificities. Out of those DP thymocytes, cells that interact at a low affinity with selfpeptide–MHC complexes displayed in the thymic cortex are induced to survive and differentiate into CD4+CD8 or CD4 CD8+ single-positive (SP) thymocytes. This process, termed positive selection, contributes to the selection of self-MHC-restricted and potentially useful T cell repertoire. Cortical thymic epithelial cells (cTECs) play an essential role in inducing the positive selection of cortical DP thymocytes [3,4]. Positively selected thymocytes are induced to express chemokine receptor CCR7 and are attracted to the thymic medulla, where CCR7 ligands are abundant [5,6]. In the thymic medulla, a variety of antigen-presenting cells (APCs), including medullary thymic epithelial cells (mTECs) and dendritic cells (DCs), present a wide range of self-antigens, including tissue-restricted self-antigens produced by mTECs, via the promiscuous gene expression mechanism. High-affinity TCR interactions with those self-antigens displayed in the thymic medulla induce the deletion of positively selected thymocytes or the differentiation into regulatory T (Treg) cells, thereby contributing to the establishment of self-tolerance in T cells [7,8]. Thus, a variety of APCs in the thymus, including cTECs, mTECs, and DCs, play important roles in the selection of developing T cells to form a TCR repertoire that is capable of responding to diverse foreign antigens and is tolerant to self-antigens. Recent studies have revealed that these thymic APCs carry unique properties in the expression, processing, and presentation of self-antigens specialized for thymocyte selection. In this brief review, we will summarize those properties unique to the thymus, with emphasis on the biology of cTECs and mTECs.

Protein degradation in cTECs for positive selection of T cells cTECs carry a number of unique machineries for protein degradation, which critically contribute to the processing of self-antigens that induce positive selection of thymocytes (Table 1). A number of recent articles discuss those machineries in detail [9,10]. Here we briefly summarize antigen-processing machineries unique in cTECs, as follows. Current Opinion in Immunology 2017, 46:53–57

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Table 1 Antigen processing and presenting machineries in thymic epithelial cells Cell cTEC

mTEC

Molecule & process

Contribution to Ag processing & presentation

Thymoproteasome Capthepsin L TSSP Constitutive autophagy CD83 & March8

MHC MHC MHC MHC MHC

class class class class class

I Ag processing II Ag processing II Ag processing II Ag processing II turnover

Aire Fezf2 Constitutive autophagy

Promiscuous gene expression Promiscuous gene expression MHC class II Ag processing

Regarding the processing of MHC class I-associated selfantigens, cTECs express a unique form of proteasome, termed thymoproteasome, which contains the cTECspecific catalytic subunit b5t (also known as Psmb11), thereby producing a cTEC-specific set of peptides in the cytoplasm (Table 1) [11,12]. Proteasome-dependent cytoplasmic peptides are transported into the lumen of the endoplasmic reticulum and are loaded onto MHC class I molecules, so that cTECs display a unique set of thymoproteasome-dependent MHC class I-associated self-peptides [12,13]. The thymoproteasome is essential for positive selection and functional fine-tuning of immunocompetent CD8+ T cells [13,14]. Thymoproteasome-dependent MHC class I-associated peptides uniquely displayed by cTECs may contribute to positive selection of CD8+ T cells, by providing structural motifs of the peptides with intermediate TCR affinity optimal for the positive selection [12,15] and/or by providing positive selection epitopes that are displayed only in the thymic cortex and that never overlap with epitopes that are displayed elsewhere in the body and that induce negative selection [16]. Regarding the processing of MHC class II-associated selfantigens, cTECs express a unique spectrum of lysosomal and endosomal proteases, including cathepsin L and thymus-specific serine protease (TSSP, also known as Prss16) (Table 1) [17–22]. These proteases are likely involved in the degradation of lyso-endosomal proteins to produce MHC class II-associated self-peptides displayed by cTECs for the optimal positive selection of CD4+ T cells [19,22]. In addition, many cTECs are constitutively active in autophagy [23–25]. As cytoplasmic components engulfed by autophagosomes are delivered to lysosomes, autophagy contributes to the processing of cytosolic proteins for presentation by MHC class II molecules [23]. Autophagy-dependent antigen processing in cTECs may be associated with the optimal positive selection of MHC class II-restricted CD4+ T cells [24,25]. Current Opinion in Immunology 2017, 46:53–57

Protein trafficking in cTECs for positive selection of T cells In addition to the unique machineries for protein degradation, cTECs are distinct in the traffic of membrane proteins, which contribute to self-antigen presentation for positive selection of thymocytes. cTECs express CD83, which interferes with the March8mediated ubiquitination and degradation of MHC class II molecules and thereby decelerates the turnover of surface MHC class II molecules expressed by cTECs [26,27]. The thymus in CD83-deficient mice is defective in the production of CD4SP thymocytes and peripheral CD4+ T cells [28], suggesting that the CD83-mediated stabilization of MHC class II molecules on the surface of cTECs critically maintains the duration of TCR interactions required for positive selection of CD4+ T cells [26,27,28]. A fraction of cTECs form multicellular complexes termed thymic nurse cells (TNCs), in which one cTEC encloses many DP thymocytes [29,30]. TNC formation appears to represent persistent interactions between cTECs and DP thymocytes, which provide a microenvironment that optimizes positive selection through secondary TCR rearrangement in DP thymocytes [31]. Indeed, cTECs are strongly adhesive to neighboring DP thymocytes and express VCAM-1, which mediates the adhesion to DP thymocytes [31,32]. Interestingly, classical analysis of MHC expression by thymocytes showed that cortical DP thymocytes are capable of acquiring MHC molecules through the transfer from non-hematopoietic cells including cTECs [33]. Thus, the highly intimate interactions between cTECs and DP thymocytes may involve the active transfer of membrane molecules, through such mechanisms as exosome transfer and trogocytosis [34– 37], which possibly play roles in facilitating antigen presentation for positive selection of T cells.

Promiscuous gene expression in mTECs for self-tolerance in T cells Positively selected thymocytes express CCR7 and migrate from the cortex to the medulla, by being attracted to CCR7 ligands abundantly produced by mTECs [5,6]. mTECs also carry a unique mechanism of gene expression, termed promiscuous gene expression (Table 1). Promiscuous gene expression enables mTECs to express virtually entire protein-coding genes in the genome, including tissue-restricted self-antigen molecules, which contribute to the establishment of self-tolerance in T cells by deleting self-reactive T cells and by promoting the generation of Treg cells [7,38]. Autoimmune regulator (Aire), a nuclear protein expressed in a fraction of mTECs, contributes to the promiscuous gene expression [39], by directly promoting gene transcription [40] and epigenetically elevating gene expression by binding to hypo-methylated lysine-4 of histone H3 [41] and by www.sciencedirect.com

Antigen processing and presentation in the thymus Kondo, Takada and Takahama 55

binding to ATF7IP–MBD1 complexes [42]. Aire deficiency in mTECs results in a defect in promiscuous gene expression, the breakdown of T cell self-tolerance, and the onset of autoimmune disease [39]. A recent study identified Fezf2, a zinc finger transcription factor, as an additional regulator of promiscuous gene expression in mTECs [43]. The spectrum of genes regulated by Fezf2 and Aire seems different, and Fezf2-expressing mTECs and Aire-expressing mTECs do not appear to overlap. Unlike Aire, whose expression in mTECs is predominantly promoted by RANK-mediated signals [44,45], Fezf2 expression in mTECs is primarily regulated by LTbR-mediated signals [43]. Nonetheless, like Aire, the lack of Fezf2 causes autoimmune disease [43].

machineries, which are crucial for the establishment of self-tolerance in T cells. Recent studies have suggested further mechanisms of the splicing of MHC class Iassociated peptides in various aspects of antigen processing, including the processing for T cell repertoire formation [53]. Further investigation of the machineries for antigen processing and presentation in the thymus should advance our understanding of the mechanisms of T cell repertoire selection.

Acknowledgements We are thankful for the research grants from MEXT-JSPS (16H02630 and 24111004 to Y. T. and 26460576 to K. T.) and the Uehara Memorial Foundation (to K. T.).

References and recommended reading Papers of particular interest, published within the period of review, have been highlighted as:

Interplay between mTECs and DCs for selftolerance in T cells mTECs are capable of directly presenting endogenous self-antigens, including promiscuously expressed selfantigens, to thymocytes for self-tolerance in both MHC class I and class II recognition specificities [8,46,47]. As in cTECs, the baseline activity of autophagy is high in many mTECs, which is involved in the negative selection of self-reactive CD4+ T cells [24,48]. In addition to the direct presentation by mTECs, mTECderived self-antigens can be indirectly presented by thymic DCs, which are predominantly located in the medullary region [8,49,50]. The ablation of MHC class II expression in hematopoietic APCs, including thymic DCs, diminishes the negative selection of CD4+ T cells that are reactive to self-antigens produced by mTECs [8]. The dynamic turnover of mTECs and the continuous supply of dying mTECs seem to contribute to the transfer of promiscuously expressed mTEC-derived self-antigens to thymic DCs [51]. Furthermore, the interaction between chemokine receptor XCR1 expressed by thymic DCs and its ligand XCL1, also known as lymphotactin, produced by mTECs in an Aire-dependent manner may enhance the proximity between thymic DCs and mTECs and the transfer of mTEC-derived self-antigens to thymic DCs [52]. Thus, mTECs directly and indirectly present self-antigens through multiple pathways. The interplay between mTECs and DCs seems important for securing self-tolerance in T cells.

Conclusions Here, we summarized unique machineries for antigen processing and presentation in thymic APCs, particularly cTECs and mTECs, which contribute to the formation of functionally competent yet self-tolerant T cell repertoire. cTECs are unique in terms of protein degradation and membrane trafficking, both of which are important for the positive selection of functionally competent T cells, whereas mTECs are unique in terms of gene expression www.sciencedirect.com

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