Antigen Presentation and T Lymphocyte Biology PDF

Slide 4 Learning Objectives – MHC & Ag Presentation 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. List and describe the structures and functions of major histocompatibility complex molecules (MHC class I & MHC class II) List the genes and describe the genetics of the major histocompatibili...

Slide 4 Learning Objectives – MHC & Ag Presentation 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. List and describe the structures and functions of major histocompatibility complex molecules (MHC class I & MHC class II) List the genes and describe the genetics of the major histocompatibility complex and other molecules involved in antigen processing and presentation; describe the nomenclature used to define MHC I and MHC-II alleles Describe the mechanisms responsible for the variability of peptides presented by MHC molecules (MHC class I & MHC class II) – Not addressed in class; refer to Characteristics of Peptide-MHC Molecule Interactions section of the book or pp. 5-6 of this handout Describe the pathways of MHC antigen processing and presentation (MHC class I & MHC class II) List and describe the cells that present antigen via MHC class I molecules as well as the cells that respond to MHC class I-restricted antigen presentation List and describe the cells that present antigen via MHC class II molecules as well as the cells that respond to MHC class II-restricted antigen presentation Describe the process (in detail) and outcomes MHC class I-restricted antigen presentation (including all described molecules, membrane-bound and soluble) Describe the process (in detail) and outcomes MHC class II-restricted antigen presentation (including all described molecules, membrane-bound and soluble) List examples of organisms whose proteins are mainly processed by the MHC class I-restricted antigen presentation machinery List examples of organisms whose proteins are mainly processed by the MHC class II-restricted antigen presentation machinery Describe the origin and source of the peptides that are presented by MHC molecules (MHC class I & MHC class II) Describe the pathway and functions of antigen cross-presentation Describe the structure and function of superantigens, how these differ from antigens, as well as the significance of superantigen activation Describe antigen presentation mechanisms involving non-classical MHC-I molecules, the nature and source of the antigens involved, as well as the cell types and receptors that interact in each case (e.g. missing-self & altered-self presentation to NK cells, CD1-mediated presentation of glycolipids to NKT cells and IELs, MHC-I-restricted antigen presentation as well as altered-self presentation to IELs) Describe antigen presentation mechanisms involving non-protein molecules such as metals Slide 5 Learning Objectives – T Lymphocyte Biology 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. List, describe, and differentiate cells of the T lymphocyte lineage (CD4, CD8, innate lymphoid cells, IELs, NK cells & NKT cells), and describe their respective functions Describe the general recirculation of T lymphocytes and explain how these cells migrate (1) to different tissues and (2) different sections of the secondary lymphoid tissues Recall the sites of T lymphocyte activation, proliferation and differentiation, and effector functions, as well as the cytokines involved in T lymphocyte activation, proliferation, and differentiation Compare and contrast T cell receptors and B cell receptors Differentiate between : TCRs and : TCRs, as well as the development and functions of the cells bearing these different TCRs Describe the : TCR complex; describe its component parts and their functions, including CD4 and CD8 co-receptor molecules, as well as the intracellular signaling events leading to T cell activation Describe the outcomes of (1) T cell activation, (2) expansion, and (3) differentiation Describe the process of T lymphocyte costimulation (including intracellular signaling events), as well as its functional importance; differentiate the outcomes of T lymphocytes that are costimulated as compared to those that are not Describe the process of clonal expansion (including the molecules that drive lymphoproliferation & associated intracellular signaling events) Recall the cytokines and surface molecules that drive the differentiation of each type of effector T lymphocyte; describe the process of T lymphocyte differentiation (including intracellular signaling events) List, contrast, and describe the effector T cell populations (TH1, TH2, TH17, CTLs, TFH & Treg), their respective functions and how they perform these functions Contrast the phenotypes of T lymphocytes (i.e. mature, naïve T lymphocytes as compared to effector T cells, as well as effector T lymphocytes among themselves) Describe how adaptive immune responses are regulated and terminated Describe the importance of memory T cells for long-term immunity; describe the phenotype of memory T lymphocytes Describe the outcomes of thymic involution and immunosenescence Slide 6 After this lecture, you should be able to at least answer the following questions… What are the functions of MHC-I and MHC-II-restricted antigen presentation? Which cells bear MHC-I molecules? Which cells bear MHC-I and MHC-II molecules? Which cells respond to (1) MHC-I-restricted antigen presentation? (2) MHC-II-restricted antigen presentation? What is the structure of the MHC-I molecule? What is the structure of the MHC-II molecule? What is the source of peptides for (1) MHC-I molecules? (2) MHC-II molecules? Where do peptides presented by MHC-I and MHC-II come from? How are proteins processed for antigen presentation? What is cross-presentation and what is its relevance? What would be the functional consequence of a missing MHC (MHC-I or MHC-II)? How are MHC molecules inherited? How many can a person have? What are super-antigens, how do they work, and what consequences are associated with super-antigens? What are the functions of non-classical MHC-I molecules? Which cell types respond to such molecules? And how do these cells respond to such stimuli? What are the properties of the different T lymphocyte subsets? What organisms are handled by different T lymphocyte subsets? What are the properties of their T cell receptors if present? And how do T lymphocytes respond to antigen presentation to effect their functions? How do T lymphocytes migrate? How and where are T lymphocytes activated (including detailed signaling) and how are T cell responses regulated and terminated? Why, where, and how do T lymphocytes commit to an effector function and what are the functions of the different effector T cell subsets? How are naIve, effector, and memory T lymphocytes differentiated? What is the function of memory T lymphocytes? Slide 7 Antigen presentation Slide 8 Overview & Structure Slide 9 Antigen Presentation – Functions Essential for: – – – – T lymphocyte development T lymphocyte activation Killing of infected & tumor cells Effector leukocyte help (e.g. macrophages, B lymphocytes) Antigen presentation to CD8 T lymphocytes: – For the ELIMINATION of INFECTED or TUMOR CELLS Antigen presentation to CD4 T lymphocytes: – For the REGULATION of the immune system (TH cells, TFH & Treg cells) Antigen presentation is involved at many stages of a T lymphocyte’s life cycle: • • • • It is required for T lymphocyte development in the thymus; It is required for its activation when fighting infection; It is required for the identification of cells that must be destroyed (e.g. infected cells or tumor cells); It is required for the activation of effector cells such as macrophages and B lymphocytes. MHC-I-restricted antigen presentation exists for the purpose of killing infected or transformed cells; cells that respond to MHC-I-restricted antigen presentation are T lymphocytes of the CD8 subset. MHC-II-restricted antigen presentation exists for the purpose of regulating immune responses; cells that respond to MHC-II-restricted antigen presentation are T lymphocytes of the CD4 subset and these (1) provide help for other leukocytes involved in an immune response (e.g. TH1, TH2, TH17, & TFH cells) or (2) prevent inflammation (e.g. Treg cells). Slide 10 Ag presentation – Functions Present Ag to T lymphocytes: – MHC-I  CD8 T lymphocytes – MHC-II  CD4 T lymphocytes .: Allow T cells to distinguish between self and non-self; MHC-I-restricted antigen presentation exists for the purpose of selecting and activating CD8 lymphocytes; MHC-II-restricted antigen presentation exists for the purpose of selecting and activating CD4 lymphocytes. This is the process that allows the adaptive immune system to distinguish between self and non-self (as well as tumor cells). Slide 11 Cells bearing MHC molecules MHC-I  Most somatic nucleated cells (APCs) MHC-II  Professional Ag presenting cells (pAPCs): • Dendritic cells • Macrophages • B lymphocytes • Thymic stromal cells MHC-I molecules are found on the surface of all somatic nucleated cells; these molecules serve to alert CD8 cytotoxic cells to kill them because they are either infected and need to be eliminated, or they are transformed and need to be eliminated (but this requires prior activation of CD8 cells by pAPCs). MHC-II molecules are usually restricted to professional antigen presenting cells (pAPCs) and serve to activate CD4 cells so that they can provide help for effector cells among others. Note that pAPCs therefore express both MHC-I and MHC-II (leukocytes are indeed somatic nucleated cells); also note, as previously seen, that antigen presentation is required during T lymphocyte development and that, therefore, thymic stromal epithelial cells are also pAPCs, but only during the process of T cell development. Slide 12 Quick assessment of what I know so far… • What is the purpose of MHC-II-restricted antigen presentation? • What cells express MHC-II molecules? • What cells respond to MHC-II-restricted antigen presentation? • What is the purpose of MHC-I-restricted antigen presentation? • What cells express MHC-I molecules? • What cells respond to MHC-I-restricted antigen presentation? Slide 13 Structure of T Cell Receptor • Two polypeptide chains,  & , of roughly equal length • Both chains have variable (V) and constant (C) regions • Genes – -chain has V & J gene segments – -chain has V, D, and J gene segments The TCR is a heterodimer composed of two glycosylated polypeptide chains belonging to the immunoglobulin superfamily, and there are two types of TCRs: α:β TCRs and γ:δ TCRs. The α:β TCR is composed of one α-chain and one β-chain, whereas the γ:δ TCR is made up of one γchain and one δ-chain. The α:β TCR is the type found on the majority of T lymphocytes, and it binds MHC/antigenic peptide complexes (MHC-I or MHC-II). Slide 14 B cell receptor vs. T cell receptor Peptide B cells directly interact with antigen T cells recognize antigens as peptides presented in MHC, i. e. are MHC-restricted… Unlike the BCR which binds free antigen, the TCR binds antigen bound to an MHC molecule, i. e. is MHC-restricted. Slide 15 Identify similarities and differences: Structure of MHC Molecules Structure of MHC class I molecules Class I MHC molecules consist of two polypeptide chains: A polymorphic α chain responsible for class I MHC specificity which is non-covalently associated with the smaller constant (i. e. nonpolymorphic) chain b2-microglobulin. Therefore, the complete class I MHC is a heterodimer. The α chain (sometimes referred to as the heavy chain) is a transmembrane polypeptide containing three domains α1, α2 and α3. The complete molecule therefore possesses four domains, the last domain being contributed by the other polypeptide chain β2-microglobulin. The peptide binding domains, referred to as the peptide-binding cleft, peptide-binding groove or peptidebinding niche, are the α1 and α2 domains of the heavy chain. Class I MHC molecules bind short peptides of 8 to 12 amino acids (generally 8-10 aa), usually by their ends but not always (anchor aa residues) – next slide. Structure of MHC class II molecules Class II MHC molecules also consist of two polypeptide chains: The α chain and the β chain; both are polymorphic. Both chains are also membrane-spanning polypeptide chains and both contain two domains: the α1 and α2 domains, as well as the β1 and β2 domains. Once again, the complete class II MHC is a heterodimer. The peptide binding domains are the α1 and β1 domains. Class II MHC molecules bind short peptides of at least 13 amino acids, and up to 21 amino acids in length with all possible lengths in between. Hence there is a greater variation in the lengths of peptides presented by MHC-II, as compared to those presented by MHC-I – next slide. Slide 16 MHC/TCR Binding Restriction MHC-I generally bind peptides 8-12 aa long MHC-II generally bind peptides 13-21 aa long Structure of MHC class I molecules The peptide binding domains, referred to as the peptide-binding cleft, peptide-binding groove or peptide-binding niche, are the α1 and α2 domains of the heavy chain. Class I MHC molecules bind short peptides of 8 to 12 amino acids (generally 8-10 aa), usually by their ends but not always (anchor aa residues). Although different class I MHC alleles bind peptides with different anchor residues, a single MHC-I allele can only bind peptides with related anchor residues (e. g. aromatic aa such as Tyr, Phe and Trp are related; other examples would be Val, Leu, Ile & Met which are all hydrophobic, or Lys & Arg which are hydrophilic). This explains why a limited number of MHC-I molecules (a maximum of 6 in the case of humans!) can present extremely high numbers of different peptides: as long as the anchor residues are present for any given MHC allele, there can be variation in the parts separating the anchor residues. The main known source of these peptides is the proteasome (or immunoproteasomes under inflammatory conditions). Structure of MHC class II molecules Class II MHC molecules also consist of two polypeptide chains: The α chain and the β chain; both are polymorphic. Both chains are also membrane-spanning polypeptide chains and both contain two domains: the α1 and α2 domains, as well as the β1 and β2 domains. Once again, the complete class II MHC is a heterodimer. The peptide binding domains are the α1 and β1 domains. Class II MHC molecules bind short peptides of at least 13 amino acids, and up to 21 amino acids in length with all possible lengths in between. Hence there is a greater variation in the lengths of peptides presented by MHC-II, as compared to those presented by MHC-I. As for MHC-I though, different class II MHC alleles bind peptides with different anchor residue preferences (but these anchor residues are located more centrally as compared to MHC-I anchor residues), but a single MHC-II allele can also bind peptides with anchor residues that are related. This again explains the variability of peptides that can be presented by only a limited set of MHC-II molecules. The source of these peptides is mainly endosomal (phagocytic/endocytic vacuoles). Slide 17 Identify similarities and differences: MHC restriction Ag presenta on is MHC-restricted: MHC restriction means that the T cell receptor must interact (“recognize”) favorably with both the antigenic peptide AND the MHC molecule (i. e. the antigenic peptide AND the 1 and 2 domains of the MHC-I molecule, or the antigenic peptide AND the 1 and 1 domains of the MHC-II molecule). Func on of MHC class I molecules The function of class I MHC molecules is the presentation of antigenic peptides to CD8+ T lymphocytes via their T cell receptor (TCR). As we will see later, binding of the TCR to the class I MHC/Ag peptide complex generally serves at first to activate CD8+ T lymphocytes, a process done by pAPCs requiring CD8+ T lymphocyte CD28 co-stimulation by pAPC CD86 (also called B7.2). This activation signals activated CD8+ T lymphocytes to then seek and destroy infected or transformed Ag presenting cells. Since CTLs are already in an activated state by that point, infected or transformed Ag presenting cells need not express co-stimulatory molecules (CD86); they only need to express MHC-I/peptide complexes. Func on of MHC class II molecules The function of class II MHC molecules is the presentation of Ag peptides to CD4+ T lymphocytes via their T cell receptor (TCR). As we will see later, binding of the TCR to the class II MHC/Ag peptide complex generally results in the commitment of naïve CD4+ T lymphocytes to a particular immune response (TH1, TH2, TH17, TFH, Treg etc.), which also depends greatly on the cytokine environment in which Ag presentation occurs as well as co-stimulation by the Ag presenting cells. As for CD8+ T lymphocyte activation, CD4+ T lymphocyte activation requires CD28 co-stimulation by CD80 (also called B7.1 present on some subsets of activated B lymphocytes) or CD86 (present on pAPCs). Slide 18 Quick assessment of what I know so far… • What is the structure of the MHC-I molecule? • What is the structure of the MHC-II molecule? • Why are structures of MHC-I and MHC-II molecules so similar? • What does MHC-restricted antigen presentation mean? Slide 19 Intro video: MHC-I: https://www.youtube.com/watch?v=VPvCekgPwRI MHC-II: https://www.youtube.com/watch?v=_8JMVq7HF2Y Antigen Processing: Origin/Source/Function Slide 20 MHC-II-restricted Ag presentation: Class II-restricted major histocompa bility complex an gen presenta on Major histocompatibility complex class II-restricted peptides are produced in acidified endocytic vesicles such as the endosomes and phagolysosomes. Within these compartments, acidic proteases, of which the cysteine proteases cathepsins H, B, D, S and L seem to play a major role, process peptides for antigen presentation. Newly synthesised, partially folded, MHC class II molecules in the endoplasmic reticulum bind to MHC class II-invariant chain (Ii) in order to prevent peptide loading inside the endoplasmic reticulum. The Ii chain then targets the MHC class II molecule to the appropriate endosomal compartment where peptide loading occurs. During endosomal trafficking [such as in the MHC class II compartment (MIIC)], the Ii chain is sequentially cleaved by cathepsins (cathepsin S in the case of antigen presenting cells), leaving only a short peptide bound to the MHC class II molecule, the class II-associated invariant chain peptide (clip). Major histocompatibility complex class II-like molecules (HLA-DM in humans for antigen presenting cells) then bind to MHC class II:clip heterodimers where they catalyse the release of the remaining clip peptide, stabilise empty MHC class II that would otherwise aggregate, and catalyse the uptake of antigenic peptides. HLA-DM proteins also serve a peptide editing function through their ability to bind and unbind peptides to and from the MHC class II molecule, in order to insure that MHC class II:antigenic peptide complexes are stable long enough to display antigen until the encounter with the appropriate CD4+ cell. Finally, the MHC class II:antigenic peptide complexes are directed toward the surface of the cell for antigen presentation. Major histocompatibility complex class II molecules bind peptides that contain thirteen or more amino acid residues, to a maximum of 21 amino acids. Slide 21 MHC-I-restricted Ag presentation & cross-presentation: Class I-restricted major histocompa bility complex an gen presenta on Antigenic peptides loaded onto MHC class I molecules are mostly generated by the proteasome. Proteasome structure and function is discussed in greater detail in the next section. Peptides produced inside the endoplasmic reticulum can be taken up readily by MHC class I molecules within the organelle, whereas peptides produced in the cytosol must first be transported into the lumen of the endoplasmic reticulum via the transporters associated with antigen processing TAP1 and TAP2. TAP1 and TAP2 are ATP-binding cassette proteins that form the transmembrane heterodimers responsible for the preferential uptake of peptides originating from the cytosol. TAP1:TAP2 heterodimers have a predilection for transporting peptides composed of eight to twelve amino acid residues possessing basic or hydrophobic residues at their carboxy-terminal end. Newly synthesized MHC class I molecules remain inside the endoplasmic reticulum until they are loaded with an antigenic peptide. This is achieved through a scaffolding of accessory and chaperone proteins. First, the emerging MHC class I  chain (also referred to as the heavy chain) binds the transmembrane chaperone protein calnexin and the thioredoxin enzyme ERp57. The function of calnexin is to keep the MHC class I  chain in an inactive, partially folded state inside the endoplasmic reticulum, whereas the function of ERp57, in association with calcineurin and calreticulin would appear to be the proper folding of the MHC class I α chain through the isomerisation of disulfide bonds present on the heavy chain. Upon binding of 2microglobulin (2m), calnexin dissociates from MHC class I  chain:ERp57:2m heteromer that now binds to a complex of proteins containing the chaperone protein calreticulin and the TAP1associated protein that serves to bridge a complex of proteins containing the chaperone protein calreticulin, an antigenic peptide, and tapasin that serves to bridge TAP1 with the MHC class I molecule and possibly stabilise the MHC class I  chain:2m heterodimer in the absence of peptide. Therefore, by analogy with the MHC class II system, tapasin may function like the invariant chain and/or HLA-DM (discussed in the next paragraph). When an appropriate peptide attaches to the binding groove of the MHC class I molecule, complete folding of the MHC molecule occurs as it is released from the calreticulin:ERp57:tapasin:TAP-1 complex. The loaded MHC class I molecule can then be transported to the cell surface, via the Golgi complex and secretory vesicles, where antigen presentation occurs following the fusion of the secretory vesicles with the plasma membrane. Cross-presenta on In some cases, material taken up through endocytosis “leaks” into the cytosol; in such cases, the “leaked” material can be processed by proteasomes and enter the MHC-I-restricted Ag presentation pathway. Therefore, cross-presenta on is the term given for extracellular material that ends up being presented by MHC-I molecules on a cell’s surface, usually a pAPC. Crosspresentation occurs when exogenous Ag “leaks” into the cytosol from endosomes by a process called retrotranslocation; in such cases, exogenous peptides leak into the endogenous pathway (Sec61 channels?) and can be displayed by MHC class I molecules. In a way, you can consider the MHC to be a very important means by which the immune system distinguishes self from non-self. The highly specific nature of the MHC-peptide-T cell receptor interactions is what allows the immune system to deal so effectively and so efficiently with pathogens. It is also the reason behind graft rejection… a topic examined in more details in the lecture entitled Transplanta on Immunology. Slide 22 Quick assessment of what I know so far… • Describe the pathway of antigen presentation for antigens taken up from the outside of the cell… • Describe the pathway of antigen presentation for antigens taken up from the inside of the cell… • Describe the pathway of antigen cross-presentation… • Where are the peptides loaded onto MHC-II molecules generated? Where do these peptides originate from? • Where are the peptides loaded onto MHC-I molecules generated? Where do these peptides originate from? • Where are the MHC molecules synthesized? • What molecule loads antigenic peptide into the peptide binding cleft for each pathway? Slide 23 Intro video: https://www.youtube.com/watch?v=ry8X5TQMZ1w&index=2&list=PLIICVGuMKtMR371pqFWqPKH_9KkK7PZo0 Antigen Presentation for T Cell Activation Requires 3 Signals Slide 24 CD4 T cell activation: *CD8 T cells require more co-stimulation than CD4 cells hence the CD4 help… Recognition of a specific peptide:MHC complex by a naïve T lymphocyte triggers signal transduction events (mediated by the cytoplasmic tails of CD3, as well as the CD4 or CD8 coreceptor) through the TCR. These events result in T cell activation which initiates a program of cell proliferation and differentiation. However, T lymphocyte activation by MHC-restricted antigen presentation alone is not sufficient to achieve effector functions; survival and differentiation signals are also required. The survival signal delivered by pAPCs is in the form of the surface co-stimulatory molecules CD80/86, also called B7.1/B7.2 in the old nomenclature; the T cell ligand for co-stimulatory molecules is CD28. Differentiation signals come in the form of cytokines, and cytokines effectively dictate the type of effector T lymphocyte (TH1, TH2, TH17, TFH, Treg) that results from antigen presentation. CD8 T cell activation requires more co-stimulation than CD4 T cell activation. Increased pAPC CD80/CD86 surface expression required for CD8 activation comes from CD4 TH1 cell help; CD40L present on the surface of activated CD4 cells (TH1) engage CD40 on the surface of the pAPC and, with cytokines such as IL-2 and IFN-, leads to high enough pAPC CD80/CD86 expression to activate CD8 cells into becoming CTLs. Slide 25 Signaling: Signal 2 – CD28 Signal 1 – TCR Signal 3 – Cytokines Ac va on – Signal 1 The :-TCR is in fact a complex of polypeptide chains: the TCR -chain, the TCR -chain, a chain, a -chain, two -chains, as well as two -chains. The :-TCR heterodimer forms the antigen recognition component of the TCR. Two heterodimers, one containing the - and chains and one the - and -chains form the CD3 signaling complex. The two -chains form a homodimer associated with the transmembrane tail of the :-TCR heterodimer. The CD3 and  homodimer cytoplasmic tails are loosely associated with a Src family tyrosine kinase, Fyn. Associated with the TCR complex is the (1) T cell co-receptor, either CD4 or CD8, both of which are tightly associated with another Src family tyrosine kinase, Lck, on their cytoplasmic domain, and (2) CD45 which is a phosphatase required for the activation of Fyn (Fyn needs to be dephosphorylated to be activated). It is estimated that 10 to 50 peptide:MHC-II complexes displayed on the surface of pAPCs is sufficient to activate CD4 T lymphocytes; CD8 T lymphocytes are estimated to require as little as 1 to 3 peptide:MHC-I complexes. When T cells are presented with sufficient amounts of peptide:MHC complexes, TCR complex clustering takes place to initiate intracellular signaling. The cytoplasmic domains of CD3 and the  homodimer contain Immunoreceptor Tyrosine-based Activation Motifs (ITAMs) that increase the efficiency of receptor signaling by creating docking sites for the kinases involved in signaling. Clustering of two or more :-TCR heterodimers with CD4 (or CD8) co-receptors and CD45 triggers the activation of Fyn when CD45 dephosphorylate it. Fyn then phosphorylates tyrosine residues in the ITAMs of CD3 and the  homodimer. As a result, ZAP-70 (-chain-associated protein kinase 70), another tyrosine kinase, binds (docks to) the phosphorylated ITAMs of the  homodimer and is in turn phosphorylated (activated) by Lck. ZAP-70 then phosphorylates the scaffold proteins LAT (Linker of Activated T cells) and SLP-76, which are linked by an adaptor protein (GADS); the LAT:SLP-76:GADS complex then recruits phospholipase C- (PLC-) which then cleaves phosphatidylinositol biphosphate (PIP2) into diacylglycerol (DAG) and inositol triphosphate (IP3). Further downstream signaling include (1) the activation and nuclear translocation of the transcription factor nuclear factor kappa B (NF-B) as a result of PKC- activation by DAG, (2) the activation and nuclear translocation of the transcription factor nuclear factor of activated T cells (NF-AT) resulting from Ca2+/calcineurin signaling by IP3, and (3) the activation and nuclear translocation of the transcription factor AP-1 (Activator Protein 1) as a result of MAPK (Mitogen-Activated Protein Kinase) activation by RasGRP and DAG (refer to the Cellular Signaling I & II lecture for a review on cell signaling). Survival and Prolifera on – Signal 2 While the ligation of the T cell receptor with specific peptide:MHC is necessary for activation, it is not sufficient. T lymphocytes also require a second signal, termed co-stimulatory signal or survival signal. The pAPC provides that signal; CD80 and/or CD86 on the surface of the pAPC bind CD28 on the surface of the T cell. Professional APCs usually do not express co-stimulatory molecules in the absence of infection; signaling through toll-like receptors or other receptors of innate immunity (inflammation) triggers the expression of co-stimulatory molecules by pAPCs. The combined peptide:MHC and CD28:CD80/86 interaction triggers complete activation of the T cell. When a naïve T cell binds to peptide:MHC on a cell that does not express CD80/86, the cell becomes non-responsive to antigen (anergic) and cannot subsequently become activated; these usually eventually die from apoptosis or become Treg cells (depending on context). CD28dependent co-stimulation drives the expression of IL-2 and high affinity IL-2 receptors (IL-2R – :: heterotrimers; naïve, resting T lymphocytes express low affinity IL-2 receptors – : heterodimers), which in turn triggers T cell proliferation and allows for differentiation to take place. CD28-mediated signaling is achieved by a variety of second messengers, including the phosphatidylinositide 3-kinase/Akt pathway, the Ras-MAPK pathway, as well as Lck. Differen a on – Signal 3 (Depends on the par cular pathogen, i. e. the cytokines provided by the innate system during Ag presenta on) Differentiation of T lymphocytes into effector cells is driven by cytokines. Generally, antigen presentation achieved in the presence of IL-12 and IFN- yields TH1 cells. Antigen presentation achieved in the presence of IL-4 (with or without combinations of IL-5, IL-10 & IL-13) generates TH2 cells. Antigen presentation achieved in the presence of IL-6, IL-23, and TGF- yields TH17 cells. Antigen presentation achieved in the presence of IL-10, TGF- or both, generates Treg cells. CD8 T lymphocytes generally differentiate into cytotoxic T lymphocytes. Cytokine signaling is primarily achieved through the Jak/STAT (Janus kinase/Signal Transducer and Activator of Transcription) signaling pathway.