a. Antigen presentation: mechanims implicated

T cell development, described in the next section, occurs in the thymus and generates a vast repertoire of naïve CD4⁺ and CD8⁺ T cells (Klein et al., 2014; Takaba and Takayanagi, 2017).

When fully mature, these cells exit the thymus, enter the circulation and can reach the secondary lymphoid organs (SLOs), among which lymph nodes (LNs), where they can be primed (Fu et al., 2016).

The specificity of T cells is encoded by their T cell receptor (TCR). In order to be activated, the TCR must recognize its cognate antigenic peptide bound to MHC molecules; MHC-I for CD8⁺ T cells and MHC-II for CD4⁺ T cells, allowing a tight control of T cell responses (Benvenuti, 2016;

Murphy et al., 2008). These peptide/MHC complexes are presented by APCs to naïve T cells, in the SLOs, structures that favour the encounter of naïve T cells with APCs having acquired antigens (Murphy et al., 2008).

Naïve T cell priming takes place in the presence of three integrated signals provided by APCs, the interface between the APC and the T cell being called the “immunological synapse” (Benvenuti, 2016). Signal 1 corresponds to the engagement of the TCR with the above-mentionned peptide/MHC complex, ensuring the specificity of the activation. Signal 2 is the ligation of co-stimulatory molecules, such as CD80/CD86 and CD40, expressed by APCs, with CD28 and CD40-L expressed by T cells, respectively. Signal 3 is the pattern of cytokines produced by APCs, which polarizes the T cell response. Signals 2 and 3 translate the state of activation of APCs by DAMPs or PAMPs, leading to the polarization of the T cell response, which is described later in this introduction (Benvenuti, 2016; Carbo et al., 2014). While signal 1 is strictly required, signal 1 in the absence of signals 2 and 3 leads to peripheral T cell tolerance (Baldwin and Hogquist, 2007;

Iberg et al., 2017).

Once naïve T cells have been primed in the SLOs, they can migrate to the tissue from which their cognate antigen has been acquired (Fu et al., 2016).

Presentation of antigens loaded onto MHC-I to CD8⁺ T cells

With the exception of erythrocytes, all cell types express MHC-I in order to present endogenous antigens, including intracellular pathogen and tumor antigens, enabling the recognition of these cells by CD8⁺ T cells in tissues, and their subsequent killing (Neefjes et al., 2011). Briefly, the classical antigen-processing pathway for the presentation of endogenous antigens to CD8⁺ T cells encompasses the degradation of antigens by the proteasome in the cytosol, and the translocation

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of generated peptides into the endoplasmic reticulum (ER) through transporter associated with antigen processing (TAP), where they are loaded onto MHC-I (Neefjes et al., 2011). Alternative antigen-processing pathways for MHC-I also exist but will not be described here (Oliveira and van Hall, 2015).

Nonetheless, APCs have the ability to internalize, process and present exogenous antigens loaded onto MHC-I to CD8⁺ T cell, a mechanism called “cross-presentation”, for which multiple pathways of antigen-processing have been described (Adiko et al., 2015; Joffre et al., 2012). In the TAP-dependent cytosolic pathways, exogenous antigens, after phagocytosis, are either exported to the cytosol, degraded by the proteasome and loaded onto MHC-I in the ER, or they are re-imported into the phagosome and loaded onto MHC-I (Gutierrez-Martinez et al., 2015; Joffre et al., 2012). Fusion between the ER and the phagosome with subsequent loading into the mix compartment has also been described (Guermonprez et al., 2003; Gutierrez-Martinez et al., 2015). The vacuolar pathway (TAP-independent) involves the degradation of antigens and the loading on MHC-I directly in the phagosome (Joffre et al., 2012).

Presentation of antigens loaded onto MHC-II to CD4⁺ T cells

In contrary to MHC-I, MHC-II expression, which is regulated by the master regulator Class II MHC complex transactivator (CIITA), is restricted to APCs (Kambayashi and Laufer, 2014;

Reith et al., 2005). The classical pathway for the presentation of exogenous antigens to CD4⁺ T cells through MHC-II, depicted in Fig. 1, involves the assembly of MHC-II molecules in the ER, where they form a complex with the invariant chain (Ii). The complex is then transported to the MHC class II compartment (MIIC), where Ii and the internalized exogenous antigens are degraded by MIIC-resident proteases (Neefjes et al., 2011; Roche and Furuta, 2015). The MHC-II peptide-binding groove is normally hidden by Class MHC-II-associated Ii peptide (CLIP), a fragment from Ii. With the help of a chaperone, called human leukocyte antigen (HLA)-DM in human and H2-M in mouse, CLIP is exchanged with the antigenic peptide, which is finally loaded onto MHC-II. MHC-II molecules are subsequently transported to the plasma membrane, where they can present the peptide to CD4⁺ T cells (Neefjes et al., 2011; Roche and Furuta, 2015).

Non-classical pathways of MHC-II loading for the presentation of endogenous antigens to CD4⁺ T cells also have been described, involving autophagy as well as non-autophagic pathways (Leung, 2015; Munz, 2015; Roche and Furuta, 2015). It is for example the case for intracellular viral antigens in infected APCs, or for the presentation of endogenously-expressed self-antigens by thymic epithelial cells (TECs), which use macroautophagy (Anderson and Takahama, 2012;

Klein et al., 2009; Veerappan Ganesan and Eisenlohr, 2017).

5 Figure 1. Classical pathway for the presentation of exogenous antigens loaded onto MHC-II to CD4⁺ T cells.

Major histocompatibility complex class II (MHC-II) is assembled in the endoplasmic reticulum (ER), where it forms a complex with Ii, the invariant chain. This complex is transported via the Golgi to the MHC-II compartment (MIIC), through the plasma membrane or directly. Ii and the exogenous antigens are degraded in MIIC by proteases. Class II-associated Ii peptide (CLIP), a fragment from Ii, hides the MHC-II peptide-binding groove, until it is exchanged with an antigenic peptide by the chaperone HLA-DM, in human (or H2-M in mice). Subsequently, MHC-II molecules are transported to the plasma membrane where they can present the peptide to CD4⁺ T cells.

APC, antigen-presenting cell; TCR, T cell receptor.

Adapted from Neefjes et al., Nat Rev Immunol, 2011 (Neefjes et al., 2011).

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