T Cell Development PDF

1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Just like the pre-BCR and BCR, the pre-TCR and TCR are inherently incapable of signaling and must therefore be associated with other polypeptide chains able to assist in signal t...

1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Just like the pre-BCR and BCR, the pre-TCR and TCR are inherently incapable of signaling and must therefore be associated with other polypeptide chains able to assist in signal transduction; six such subunits, collectively referred to as CD3, cluster with the TCR to provide phosphorylation sites (ζ[zeta]-chains) on the cytoplasmic side of the plasma membrane for the recruitment the next kinase in the signaling cascade. Thymocytes start to rearrange their β-, δ-, and γ-chain genes at the same time in a competitive manner. By contrast, in B cells, immunoglobulin gene rearrangement is orderly and sequential. The appearance of one set of receptors signals the cessation of rearrangement of the competing set of Top receptors. If a functional γ:δ receptor is produced before the elaboration of a functional βchain, then a γ:δ TCR-expressing T cell develops (differences between α:ß and γ:δ T cell populations will be discussed below) and exits the thymus without undergoing any selection process. In thymocytes that express a functional b-chain first (the majority), the ß-chain https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 20/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 associates with an invariant pre-T α-chain (a surrogate α-chain expressed until the formation of a bone fide α-chain) and the CD3 molecule (see below for a description of CD3), forming a pre-TCR complex. Expression of the pre-TCR signals double-negative thymocytes to stop rearrangements of the b-chain gene and undergo proliferation. The cells are also signaled to initiate expression of CD4 and CD8, giving rise to double-positive (DP) thymocytes (CD4+CD8+). CD4 is a molecule composed of a single polypeptide chain of the immunoglobulin superfamily. CD8, on the other hand, is composed of a heterodimer made up of an a-chain and a b-chain; both polypeptide chains also belong to the immunoglobulin superfamily. Once they complete proliferation, rearrangement of the TCR a-chain ensues. While this is occurring, γ - and δ-chain rearrangements are ongoing and, again, if a functional γ:δ TCR is produced first, the cell commits to the γ:δ lineage and exits the thymus; if on the other hand, a functional α:ß TCR is produced first, the cell commits to the α:ß lineage and further thymic development will continue. The expression of the TCR signals the end of all gene rearrangements and stimulates the expression of other T cell markers (see below). These cells have a lifespan of about 3-4 days and die of apoptosis unless they are rescued by engagement of their TCR (positive selection – see below). Only 10-30% of TCRs generated (i.e., 10-30% of DP cells) will interact with self-peptide:self-MHC complexes. Cells that fail to express a TCR die by apoptosis and are phagocytosed by macrophages in the thymic cortex. Which of the following enzymes is involved in the rearrangement of the TCR?  Autoimmune regulator (AIRE)  Stem cell factor (SCF)  Recombination-activating genes 1 & 2 (RAG1/RAG2)  Activation-induced cytidine deaminase (AID)  Adenosine deaminase (ADA)  Check Top Which of the following enzymes is involved in the rearrangement of the TCR? Adenosine deaminase (ADA) Activation-induced cytidine deaminase (AID) https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 21/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Autoimmune regulator (AIRE) Recombination-activating genes 1 & 2 (RAG1/RAG2) (Correct answer) Stem cell factor (SCF) Top The TCR complex (TCR/CD3 complex) is invariably associated with a co-receptor (CD4 or CD8) that (1) binds the MCH, and (2) possesses, associated with its cytoplasmic tail, the tyrosine kinase (primarily a TK called lymphocyte-specific protein tyrosine kinase, or lck) responsible for the phosphorylation of CD3 ζ-chains and the next tyrosine kinase in the pathway (i.e., ZAP-70) https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 22/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 (zeta-chain-associated protein kinase; this will be further developed in the Antigen Presentation and T Lymphocyte Biology activity). Like B cell receptors, T cell receptors must undergo a selection process. T cell receptors must be tested to (1) ensure the TCR can interact with self-MHC (positive selection) and (2) that the TCR does not respond to self-antigen (negative selection), i.e., is autoreactive with the potential to cause autoimmune disease. There are two positive/negative selection pathways: one is sequential, and the other is nonconsecutive. Click each tab below to learn more about the different pathways. Sequential Pathway Nonconsecutive pathway In the sequential pathway, thymocytes whose TCR binds self-MHC:self-peptide with moderate affinity are given survival signals and are thereby positively selected. Those that fail to sufficiently interact with self-MHC die by neglect and are thereby negatively selected. Positive selection occurs deep in the thymic cortex, and the antigen-presenting cells involved in the process of positive selection are cTECs; these cells present tissue-restricted peptides in the context of both MHC class I and MHC class II. Thymocytes positively selected for MHCreactivity in this way must then be tested for self-reactivity, which mostly occurs in the medulla: TCRs that bind self-MHC:self-peptide too strongly are signaled to die, whereas the others survive, mature, and exit the thymus. Positive selection leads to the maturation of cells that can sufficiently react with the person’s MHC allotypes and also determines if a double-positive cell becomes a CD4+ cell or a CD8+ cell. Double-positive cells interact with self-peptide: selfMHC complexes. When a TCR preferentially interacts with a class I MHC molecule, the cell commits to the CD8+ cell lineage; likewise, when a TCR preferentially interacts with class II MHC, the cell commits to the CD4+ lineage. The CD4 and CD8 co-receptor molecules then become part of the TCR complex. Cells that remain CD4+CD8+ are generally negatively selected – in other words, if the TCR fails to interact with either MHC class I or MHC class II, the cell dies. Thymocytes that have been positively selected move on to the thymic medulla, where they are presented self-peptides by medullary thymic epithelial cells (mTECs), bone marrow-derived dendritic cells (bmDCs), and bone marrow-derived macrophages; these cells express tissue-specific antigens (syn. tissue-restricted antigens), as a result of so-called promiscuous gene expression, that would otherwise never be expressed in the thymus (e.g., insulin and hundreds of tissue-specific proteins) and present these tissue-specific peptides to thymocytes in an effort to eliminate thymocytes reactive to these tissue-specific proteins. This Top is possible due to a singular transcription factor these cells express, an autoimmune regulator, or AIRE, which activates these tissue-specific genes. https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 23/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Watch this video: AIRE: From promiscuous molecular partnerships to promiscuous gene expression (https://www.youtube.com/watch? v=lUYeMQmGmO4) One key factor in the process of negative selection is the ability of these cells to provide costimulation (CD80/CD86 – see below in T Lymphocyte Signaling); thymocytes specific for selfMHC: self-peptide complexes that receive co-stimulation in the thymic medulla are signaled to die, thereby eliminating those positively selected thymocytes that are self-specific and could potentially lead to autoimmune disease. Note that, as we will see later, once in the periphery, antigen presentation with co-stimulation yields activation, clonal expansion, and differentiation, not death. After the negative selection process, thymocytes that have escaped negative selection achieve maturation and leave the thymus as mature, naïve (i.e., have never been presented with foreign Ag), resting α:ß T lymphocytes. Overall, T lymphocyte development takes place in about 3 weeks, and at the end of the process, only about 2% of DP cells survive the selection process and are exported to the periphery. Negative selection is the main driving force behind the development of central tolerance (i.e., the process by which one’s immune system tolerates one’s antigens by eliminating the most self-reactive T lymphocytes in the thymus). Tolerance, as well as the breakdown of tolerance, is an important concept when considering autoimmune disease, hypersensitivities, and tissue transplant. In the nonconsecutive pathway, thymocytes with high TCR affinity are signaled to die and deleted (central tolerance), whereas those with moderate TCR affinity go on to become mature, naïve T lymphocytes; thymocytes with intermediate TCR affinity develop into thymic regulatory T lymphocytes (tTreg) that act as suppressive cells in the periphery to repress immune reactivity to self-antigen previously seen in the thymus and thus contribute to peripheral tolerance. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 24/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 What process leads to the elimination of self-reacting (self-specific) lymphocytes?  Single-positive lineage commitment  Positive selection  Peripheral tolerance  Clonal expansion  Negative selection Top  Check https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development  25/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Question 1 What process leads to the elimination of self-reacting (self-specific) lymphocytes? Positive selection Peripheral tolerance Clonal expansion Negative selection (Correct answer) Single-positive lineage commitment Question 2 Which of the following describes the elimination of self-reacting T lymphocytes in the thymus? Positive selection Peripheral tolerance Central tolerance (Correct answer) Immune senescence Clonal expansion Mature T cells exit the thymus and enter the pool of recirculating lymphocytes. Mature, naïve T lymphocytes that encounter large, constant amounts of antigen shortly after exiting the thymus and in specific locations (e.g., self-antigen for those few T cells that have eluded negative selection, food, normal microbiota, dust, pollen etc.) are either eliminated by activation (activation-induced cell death), become anergic due to lack of co-stimulation (pAPCs presenting these antigens usually do so in absence of co-stimulation – many of these cells can survive in an anergic), or are suppressed by anti-inflammatory cytokine stimulation expressed by Treg lymphocytes. Eliminating or rendering T lymphocytes unresponsive to ‘ubiquitous’ antigens once they have left the thymus and reached the periphery is called peripheral tolerance. Mature, naïve, resting T lymphocytes are longer-lived than B cells and, in the absence of antigen stimulation, may live for years. In the presence of antigens, they may become activated and release cytokines that mediate effector functions (these cells have a dramatically shortened lifespan – days to weeks) or become memory cells responsible for long-term immunity (with a lifespan of months to years – see below). Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 26/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Which of the following describes the regulation of self-reacting T lymphocytes in the periphery?  Central tolerance  Negative selection  Immune senescence  Peripheral tolerance  Positive selection  Check Which of the following describes the regulation of self-reacting T lymphocytes in the periphery? Negative selection Positive selection Peripheral tolerance (Correct answer) Central tolerance Immune senescence  To perform their function(s), T lymphocytes require three separate signals: (1) activation per se (i.e., TCR complex engagement), (2) co-stimulation (i.e., CD28 engagement to reinforce signal 1), and (3) differentiation (i.e., cytokine receptor(s) engagement); signal 2 is required to reinforce signal 1 and make the activation complete (survival and clonal expansion of the lymphocytes), whereas signal 3 is the signal that actually determines the subset of T lymphocyte the cell will differentiate into (e.g., T lymphocyte required for an immune response to intracellular microbes, as opposed to an immune response against extracellular microbes, each of which requiring different Top strategies). https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 27/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 To perform their function(s), T lymphocytes require three separate signals: (1) activation (i.e., TCR complex engagement), (2) co-stimulation (i.e., CD28 engagement to reinforce signal 1), and (3) differentiation (i.e., cytokine receptor(s) engagement). Recognition of a specific peptide: MHC complex by a T lymphocyte triggers signal transduction events mediated by the TCR complex. As previously mentioned, the TCR complex is composed of two parts: (1) the TCR chains, which are responsible for Ag recognition and, therefore, antigen specificity, and (2) a set of transmembrane molecules (CD3 as well as CD4 or CD8 depending on the T lymphocyte subset) responsible for the signal transduction leading to physiological responses. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 28/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 The TCR α- and β-chain extracellular domains provide the specificity of the TCR complex, whereas CD3 provides the signaling capacity of the TCR complex. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 29/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 What part of the TCR complex allows it to transduce antigen recognition into an intracellular messaging response?  CD28  CD3  CD80/CD86  CD1  TCR α- and β-chains  Check What part of the TCR complex allows it to transduce antigen recognition into an intracellular messaging response? CD1 CD3 (Correct answer) CD28 CD80/CD86 TCR α- and β-chains However, T lymphocyte activation by MHC-restricted antigen presentation alone is not sufficient to achieve a physiological response; survival and differentiation signals are also required. The survival signal is delivered by pAPCs in the form of the surface co-stimulatory molecules CD80 and/or CD86, also called B7.1 and B7.2, respectively (or B7 in the old nomenclature); the T cell surface receptor for co-stimulatory molecules is CD28. However, co-stimulation has different consequences depending on the location where Ag presentation occurs. During negative selection in the thymus, Ag presentation and co-stimulation leads to T cell death and, therefore, is the process driving central tolerance. In peripheral (secondary) lymphoid tissue, activation of mature T lymphocytes also requires differentiation signals in the form of cytokines, and these cytokines effectively dictate the type of effector T lymphocyte subset (TH1, TH2, TH17, TFH, Treg) that results from antigen presentation. So, in summary, Ag presentation and CD80/CD86 co-stimulation in the Top is responsible for central tolerance, whereas Ag presentation thymus results in T cell apoptosis and in peripheral lymphoid tissue requires survival and proliferation signals (CD80/CD86) as well as cytokine signaling to differentiate T cells according to the immune functions required. Finally, incomplete T lymphocyte activation in secondary lymphoid tissue, as previously mentioned, leads https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 30/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 to T cell anergy and peripheral tolerance. Note: T lymphocyte signaling will be further developed, in detail, in the Antigen Presentation & T Lymphocyte Biology lecture. Top T cells failing to receive co-stimulation during antigen presentation either become (1) anergic (non-responsive to antigenic stimulation) or (2) become tolerant to said antigen. https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 31/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 What is the T lymphocyte receptor for costimulatory molecules?  CD80/CD86  CD28  CD1  CD3  TCR α- and β-chains  Check What is the T lymphocyte receptor for costimulatory molecules? CD1 CD3 CD28 (Correct answer) CD80/CD86 TCR α- and β-chains As aforementioned, the differentiation of T lymphocytes into effector cells in peripheral lymphoid tissue is driven by cytokines. Generally, antigen presentation achieved in the presence of IL-12 and IFN-γ yields TH1 cells; TH1 cells are involved in adaptive immune responses to intracellular pathogens (viruses and intracellular bacteria, fungi, & protozoans). Antigen presentation achieved in the presence of IL-4 (with or without combinations of IL-5, IL-10, & IL-13) generates TH2 cells; TH2 cells are involved in adaptive immune responses to helminths (worms). Antigen presentation achieved in the presence of IL-1ß, IL-6, IL-23, and TGF-ß yields TH17 cells; TH17 cells are involved in adaptive immune responses to microscopic extracellular pathogens (some bacteria, fungi, & protozoans) but also in inflammatory and autoimmune diseases; TH17 cells activated in the presence of IL-6 and TGF-β yield so-called inflammatory TH17 cells, also called regTH17 cells Top extracellular microbes), whereas TH17 cells activated in the (these are the cells useful to eliminate presence of IL-1β and IL-23 yield so-called pathological TH17 cells (those involved in inflammatory and autoimmune diseases). Antigen presentation achieved in the presence of TGF-ß generates https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 32/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Treg cells. CD8 T lymphocytes generally differentiate into CTLs (cells that are specialized in the killing of infected host cells and transformed cells. Ag presentation is a part of several stages of the T cell’s life cycle, and at each stage, Ag presentation serves a different function. Click through this interactive to learn more about Ag presentation. Ag Presentation in the Life Cycle of a T Cell START Top Process Site https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 33/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Antigen presentation for the purpose of T lymphocyte development (Positive and negative selection) Antigen presentation to double-positive thymocytes by thymic cortical epithelial cells (positive selection) and bone marrow-derived macrophages and dendritic cells (negative selection - i.e., central tolerance) Thymus - Development (Primary lymphoid organ) Antigen presentation for the purpose of T lymphocyte development (Response to infection) Antigen presentation to naïve T lymphocytes by professional antigenpresenting cells (mostly dendritic cells & macrophages) Lymph nodes, spleen - Activation (Secondary lymphoid organs) Requires co-stimulatory signals (CD80/CD86) Antigen presentation for the purpose of effector responses (B lymphocyte help, macrophage activation, etc.) Site of infection & secondary follicules HELP (Affected tissue) Antigen presentation to T helper (TH) lymphocytes (activated T cells) at site of infection so that TH cells can deliver further signals to heighten the effector responses of the cells presenting antigen (killing of microbes, antibody production, etc.) No requirement for co-stimulation Antigen presentation for the purpose of reactivation of memory T lymphocytes Top Antigen presentation to memory T lymphocytes upon re-exposure to infection to activate the anamnestic response https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 34/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 We will not go into future detail on this topic.  Depending on the cytokine(s) providing the differentiation signal(s) during antigen presentation and co-stimulation, T lymphocytes differentiate into subsets specialized into responding to specific types of infections. Top As mentioned above, there are two forms of TCRs: α:β TCRs present on the majority of T cells, and γ:δ TCRs present on some T cell subsets. T lymphocytes that express α:ß TCRs recognize peptides bound to MHC (refer to MHC & Antigen Presentation handout material). As described https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 35/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 above, α:ß TCRs that preferentially interact with peptides presented by MHC class I also express the CD8 co-receptor, while those that preferentially interact with peptides presented by MHC class II express the CD4 co-receptor. The activation of CD8+ T lymphocytes generally yields cytotoxic T lymphocytes (C TLs) and memory CTLs. The activation of CD4+ T lymphocytes yields distinct T lymphocyte effector and memory cells: T helper 1 (TH1), T helper 2 (TH2), T helper 17 (TH17, also called regulatory TH17 cells or Treg17), T follicular helper (TFH), regulatory T (Treg) cells (discussed in the next section), and many other T lymphocyte subsets we will not discuss. γ:δ TCRexpressing T cells comprise a small population of the T cell pool (<5%). γ:δ T cells can be CD4+, CD8+, or CD4-CD8- and recognize peptide and non-peptide antigens (e.g., bacterial cell wall phospholipids or bacterial glycolipids) in the presence or absence of MHC (refer to MHC & Antigen Presentation handout material and in a later section). They appear to recognize commonly occurring microbial components and contribute to the body’s first line of defense (i.e., act as part of the innate system; hence they are referred to as innate-like lymphocytes (ILLs), which include intraepithelial lymphocytes (IELs), natural killer cells (NK cells), and natural killer T lymphocytes (NKT lymphocytes); as opposed to IELs and NKT lymphocytes, NK cells do not possess a TCR however. Intraepithelial lymphocytes (IELs) are a heterogeneous group of cells that are found in the epithelium (about one IEL for 10-15 epithelial cells); 90% of these cells are T lymphocytes, and 80% of them carry CD8 and although they can have either α:α, α:ß or γ:δ TCRs, most of them express γ:δ TCRs – so we generalize that intraepithelial cells are γ:δ CD8 T lymphocytes; in contrast to the CD8 T lymphocytes you are accustomed to, these IELs do not require prior MHC-Irestricted activation by pAPCs to kill their cellular target and, therefore, are considered to be part of the innate compartment; also, they are unusual in that they kill infected cells either through MHC-Irestricted Ag presentation, just like CTLs, or through MHC-I-like molecule stimulation expressed by stressed or infected cells, just like NK cells; in a nutshell, you could think of IELs as a kind of innate CTL-NK cell hybrid! Intraepithelial cells fall into two classes: Type a IELs and Type b IELs. Type a IELs are IELs composed of T lymphocytes that follow the conventional development of T lymphocytes in the thymus (i.e., they go through both positive and negative selection), and the vast majority of them are CD8+ T lymphocytes. These possess a conventional α:ßTCR, as well as a conventional α:ß heterodimer CD8 co-receptor. They are activated in Top MALTs and the skin. Since these cells possess a conventional α:ßTCR and a conventional a:b heterodimer CD8 co-receptor, they kill cells in an MHC-I-restricted manner the way CTLs do. Also, since they undergo negative selection in the thymus, few self-reactive IELs leave the thymus. Type a IELs differ from CTLs in that they express high levels of NKG2D https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 36/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 homodimers (also found on the surface of NK cells), a receptor that activates killing by binding to non-classical MHC-I molecules (e.g., MICA & MICB). Type b IELs, on the other hand, are unconventional CD8+ T lymphocytes in that they possess an α:αCD8 homodimer co-receptor instead of the conventional α:β heterodimer CD8 co-receptor. This α:α CD8 homodimer co-receptor is paired with either a conventional α:ßTCR or α γ:δTCR. However, Type ß IEL α:ßTCRs or γ:δTCRs do not bind conventional peptide: MHC-I ligands; instead, they bind non-classical MHC-I molecules and other ligands (e.g., PAMPs). Since these cells do not interact with conventional peptide: MHC-I ligands and they possess an unconventional α:αCD8 homodimer co-receptor, there is little potential for autoimmunity; unconventional α:αCD8 homodimer co-receptors have very low affinity for conventional peptide: MHC-I ligands. Type ß IELs, like Type α IELs, possess high levels of NKG2D homodimers, which can also activate killing. The development of Type ß IELs is poorly understood. Unconventional CD8+ T lymphocytes may arise from late DN/early DP thymocytes that partially undergo positive selection in the thymus but escape negative selection due to the low-affinity α:α CD8 homodimer co-receptor. They then exit the thymus to finalize their maturation in mucosal epithelia or the skin, a process that seems to also require IL-15. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 37/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 1. The virus invades a cell in the mucosal epithelium. 2. The cell that has been infected presents viral peptides to CD8 IEL through MHC class 1 molecules. 3. The activated IEL eliminates the infected epithelial cell through perforin/granzyme and the Fas-dependent pathways. 4. Infection, damage, or exposure to toxic peptides induces stress in epithelial cells, leading to expression of MIC-A and MIC-B. 5. The NKG2D receptors on IELs engage with MIC-A,B, triggering IEL activation. Simultaneously, CD8 homodimers interact with TL molecules. 6. The stressed cell is targeted for destruction by the activated IEL through the perforin/granzyme pathway. Intraepithelial cells are innate-like lymphocytes found in mucous membranes and can innately recognize and kill infected cells the same way NK cells and cytotoxic CD8 lymphocytes do. Regulatory T lymphocytes (Treg) express the α:ß-TCR, as well as the CD4 co-receptor and high levels of CD25 (a component of the receptor for IL-2, the IL-2 R α-chain) and CD152 (CTLA-4 – a negative regulator of T cell activation). Despite positive and negative selection, a few self-reactive T cells are maintained and allowed to exit to the periphery. Indeed, in order to be able to deal with microbial antigens that may closely resemble human antigens, some T cells with potential selfreactivity must be allowed to escape negative selection. Treg cells exist to suppress the response of self-reactive CD4 T cells; they are referred to as natural regulatory T lymphocytes and are important in exerting peripheral tolerance. They are characterized by the expression of CD25 and CD152 on their cell surface and by the activation of the transcriptional repressor protein FoxP3. The exact mechanism by which these cells exert their effects is not completely understood. It is clear, though, that these cells play an important role in regulating normal immune cell functions; individuals (males, as FoxP3 is encoded on the X chromosome) with FoxP3 deficiency develop fatal autoimmune disorders characterized by autoimmunity against a variety of tissues. Other T lymphocyte subsets exist but will not be developed further (this being said, some of the following subsets are going to be tested on NBME/USMLE tests at some point in the near future, but I will keep track and determine when to further develop these.). These include (1) TH3 lymphocytes, mostly another type of regulatory T lymphocyte, and function to control immune responses at the mucosa level, (2) TH22 (CD4+ IL-22 secreting cells) and cytotoxic (3) Tc22 lymphocytes (cytotoxic CD8+ cells that also secrete IL-22) cells that are involved in inflammation and wound healing, (4) TH9 lymphocytes Top (IL-9-secreting cells) involved in antitumor immunity as well as both inflammatory and regulatory processes – and involved in autoimmune disease and allergy, (5) CD8+ inducible Treg cells, (6) pathogenic TH17 cells, (7) T H1/H17, and others. Other lymphoid cells bearing similarities with T lymphocytes include natural killer cells (NK cells) and natural killer T lymphocytes (NKT cells). These cells respond to MHC-I-like proteins (refer to https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 38/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 the MHC & Antigen Presentation handout). Such proteins include HLA-E (human leukocyte antigen E), MICA (MHC-I chain-related gene A), and MICB (MHC-I chain-related gene B), which are induced by cellular stress to signal cytotoxic cells that stressed cells need to be eliminated. For example, binding of NKG2D homodimers, a KAR (killer activating receptor) of NK cells, to MICA or MICB on the surface of a stressed or infected cell leads to the destruction of the cell by NK cells, provided that the stressed cell expresses little or no MHC-I. Natural killer cells will be dealt with in the Innate Immunity lecture and will no longer be developed here. The NKG2D ligands consist of MHC-like molecules, MIC-A, MIC-B, RAET1 family members, and their expression is provoked by cellular stress. Infected and transformed cells express ligands (MIC-A & MIC-B (MHC class I polypeptide–related sequencesTop A & B, & UL16 binding protein 2 (ULBP2)) that engage natural killer cell killer-activation receptors (KARs, i.e. NKG2D); some of these ligands are encoded in the MHC locus (e.g. MIC-A & MIC- B) and are referred to as modifiedself molecules, i.e. indicating the cell is altered beyond repair and needs to be killed. https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 39/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 A special case of antigen presentation is that of the NKG2 family of receptors found on the surface of NK cells and IELs. NKG2-mediated cell killing often relies on binding to nonclassical MHC-I molecules (syn. modified-self molecules) that are usually only expressed on stressed, infected or transformed cells; such non-classical MHC-I molecules include MICA, MICB and HLA-E. For example, NKG2D homodimers, one of the most common KARs present on the surface of NK cells, bind MICA or MICB; binding of MICA or MICB to NKG2D homodimers signals the NK cell to kill the cell it has docked to. This works because, usually, cells that express non-classical MHC-I proteins down-regulate their expression of MHC-I to minimize killer immunoglobulin-like receptor (KIR) signaling. Other KARs on the surface of NK cells include NKG2A/NKG2C heterodimers as well as CD94/NKG2A, CD94/NKG2B, CD94/NKG2C heterodimers, and these all bind the nonclassical MHC-I HLA-E. So these are all cases where NK cell KARs bind what are referred to as modified-self proteins (i.e., MICA, MICB, and HLA-E are considered modified-self proteins). Lastly, another example of KAR, which only recognizes pathogen determinants (PAMPs, not modified-self proteins), is NKp46; this KAR binds viral protein and heparan sulfate proteoglycans embedded in cells. Drag the receptor to the respective ligands in which it best fits. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 40/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 KIR KAR HLA-E MHC-I PAMPs MICA MICB  Check KIR MHC-I KAR PAMPs MICA MICB HLA-E Top (CD1a, CD1b, CD1c, CD1d, & CD1e) present glycolipid Another group of MHC-I-like molecules antigens to NKT cells. These cells share properties common to both T lymphocytes (CD3+ & α:ß TCR, but that can either be CD4+CD8-, CD4-CD8- or CD4-CD8+) and NK cells (NK1.1). Glycolipid antigen presentation in this manner mainly leads NKT cells to secrete cytokines (e.g., IL-4 or IFNhttps://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 41/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 γ) in a context-dependent manner (i.e., depending on the pathogen) some NKT cell subsets can also be cytotoxic and destroy infected cells. Are different from NK cells and T lymphocytes Are generally considered innate-like lymphocytes Are a diverse set of T lymphocytes that express both T lymphocyte markers (CD3+, CD4+CD8-, CD4-CD8-, CD4-CD8+, α:ß TCR) and NK cell markers (NK1.1+) Two groups of CD1: Group 1: CD1a, CD1b, and CD1c: bind glycolipids, phospholipids, and lipopeptides derived from microbes, such as mycolic acid, lipoarabinomannan, glucose monomycolate, phosphoinositol mannosides and isoprenoid glycolipids of mycobacteria Group 2: CD1d: thought to mainly bind self-lipid Ag such as sphingolipids and diacylglycerols CD1e is considered an intermediate between groups 1 and 2 React to glycolipid Ag presented by MHC-I-like molecules (CD1a, CD1b, CD1c, CD1d & CD1e), which are structurally similar to MHC-I molecules, associated with b2m, but have deeper binding grooves to accommodate hydrocarbon chains of glycolipids (hydrophobic groove binds alkyl chains to allow exposure of variable carbohydrate head and other hydrophilic groups to TCR) Secrete cytokines (e.g., IL-4, IFN-γ) in a context-dependent manner; some subsets can be cytotoxic Are important in the innate control of some infections and help direct adaptive immune responses (e.g., recognition of mycobacteria) Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 42/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 What is the ligand for the NKT lymphocyte TCR?  MHC-I/phospholipid complex  MHC-I/peptide complex  CD1/peptide complex  MHC-II/lipopeptide complex  CD1/glycolipid complex  Check What is the ligand for the NKT lymphocyte TCR? CD1/glycolipid complex (Correct answer) CD1/peptide complex MHC-I/phospholipid complex MHC-I/peptide complex MHC-II/lipopeptide complex  Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 43/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Full activation of T lymphocytes yields thousands and thousands of effector and memory T cells; effector T lymphocytes are those involved in the ongoing immune response and die shortly after activation (e.g., T helper cells or cytotoxic T cells), whereas memory cells are long-lived (months to years and decades) set aside in case of any future re-exposure to the same antigen (infection). The memory response is not only produced by circulating antibodies but also by long-lived memory B cells and memory T cells. These memory cells are generated during the primary immune response against the pathogen from a subset of effector cells or directly following peripheral activation. While most effector cells are short-lived, these memory cells survive throughout the course of the primary immune response; once the infection is resolved, memory cells proliferate and go on to survive for long periods of time – some throughout the life of the individual in the case of memory T lymphocytes. After the primary adaptive immune response, pathogen-specific memory cells outnumber their naïve counterparts by several orders of magnitude. Memory cells have the capacity to respond to specific antigen faster and better than naïve cells. While the molecular and cellular interactions necessary for Top the activation of memory cells are similar to naïve cells, memory cells are more sensitive to infection, more easily activated, and more abundant (10 – 1,000 fold) than primary naïve lymphocytes specific for the same antigen. There exist different subsets of memory T cells (e.g., central memory T cells or TCM, peripheral memory T cells or TPM, tissueresident T cells or TRM, & virtual memory T cells or TVM) and, depending on the subset, these can https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 44/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 be found in peripheral lymphoid tissue, the circulation, as well as different other tissues. For example, memory B cells, in addition to being more numerous than their naïve counterparts, have already undergone isotype switching and somatic hypermutation and, as a consequence, produce antibodies with (1) higher affinity (affinity maturation) and (2) the appropriate isotype for the pathogen, as compared to an antibody produced during the primary response. Also, upon subsequent infection, memory cell activation inhibits the activation of their naïve counterparts so that responses are skewed toward the activation of the most efficient cells. As previously mentioned, memory T cells specific for a particular pathogen are present in much higher numbers than their naïve counterparts, and therefore a greater expansion of specific T cells can occur during the secondary immune response. Additionally, memory T cells express a different profile of cell-surface molecules that allow them to perform their function and enter the peripheral tissues faster. Unfortunately, the processes involved in memory T lymphocyte development are poorly understood. The best-characterized T cell memory development is that of CD8 T lymphocytes. Memory CD8 T lymphocyte development relies on CD4 T lymphocyte help and IL-2 stimulation since CD8 T lymphocytes that do not express CD40 cannot become memory T cells, whereas those that express CD40 can. Likewise, CD8 T lymphocytes that do not express the a-subunit of the IL-2 receptor cannot become memory T cells, whereas those that express it can. A marker of memory T cells is CD45RO (as opposed to CD45RA on naïve T cells). Top Memory lymphocytes are responsible for the faster and more intense responses to subsequent antigen exposure (anamnestic response). https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 45/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 What cell type is responsible for the anamnestic response?  Neutrophil  DC  cTEC  Memory T lymphocyte  Macrophage  Check What cell type is responsible for the anamnestic response? cTEC DC Macrophage Memory T lymphocyte (Correct answer) Neutrophil  Thymus development occurs before birth and the thymus begins degenerating one year after birth, replaced by fat tissue. Thymic degeneration progresses steadily through life (thymic involution) and, as a result, a reduced production of new T lymphocytes occurs, but this does not impair T cell immunity (neither does thymectomy in adults) because T lymphocytes are longed-lived, selfrenewing, and because of memory T cell expansion upon antigen exposure – by adulthood, Top individuals possess memory T lymphocytes to the most common potentially life-threatening pathogens (childhood infections), thereby ensuring good immune protection throughout life, although T cell protection starts waning in seniors. https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 46/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 With aging and thymic involution, immunity is maintained by memory cells. Complete the activity below. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 47/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Associate the statements that best correlate. Mostly naïve T lymphocyte repertoire Children, adolescents, & young adults Older adults & elderly Mostly memory T lymphocyte repertoire Mostly naïve T lymphocyte repertoire Mostly memory T lymphocyte repertoire Children, adolescents, & young adults Older adults & elderly During differentiation, some activated T cells acquire the capacity to express the proteins they will need to perform their effector functions and others become memory cells set aside for possible future infections. Click through this interactive to learn more about T cell differentiation. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 48/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 T Cell Differentiation START Location Periphery Stage Naïve, resting T lymphocyte (TH0- Longlived) Secondary lymphoid tissue TH 0 TH 1 TFH Secondary lymphoid tissue and Action Re-circulation between secondary lymphoid tissues Antigen presentation, activation, proliferation, and differentiation into either TH2 TH17 TregTop Effector functions (Short-lived) CD8 CTL Memory cells (Longlived) https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 49/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 site of infection Death Reactivation upon Ag presentation  Work through these questions to prepare for the in-class activity. 1. Describe and contrast α:ß T lymphocytes and γ:δ T lymphocytes. 2. Describe the process and function of positive selection. 3. Describe the process and function of negative selection. 4. Describe the process of T cell activation. 5. List T helper subsets and which subset is specific to which type of pathogen.  Quiz | T Lymphocyte Development (https://rossmed.instructure.com/courses/3318/quizzes/19384)  Dr. Marc Bergeron Email: [email protected] (mailto:[email protected]) Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 50/50