1. ANTIGEN PROCESSING
and
PRESENTATION OF CELLS
Alric V. Mondragon, MD
Section of Allergy and Immunology
University of the Philippines – Philippine General Hospital
2. Outline
I. Properties of Antigens Recognized by T
Lymphocytes
II. Antigen Capture and the Functions of Antigen-
Presenting Cells
III. Processing of Protein Antigens
IV. Presentation of Non-protein Antigens to Subsets
of T Cells
4. T Lymphocytes
• Principal functions of T lymphocytes
a. to eradicate infections by intracellular microbes
b. to activate other cells, such as macrophages and
B lymphocytes.
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
5. T Lymphocytes
• Several challenges to T cells:
1. Very few naive T cells specific for any one antigen
• APCs
2. Most T cell functions require that they interact with
other cells.
• MHC
3. Different T cells have to be able to respond to
microbial antigens in different cellular
compartments.
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
6. PROPERTIES OF ANTIGENS
RECOGNIZED BY T LYMPHOCYTES
1. Most T Lymphocytes recognize only short peptides
– Induced by foreign protein antigens or small chemical
substances
2. Antigen Receptors of CD4+ and CD8+ T cells are
specific for peptide antigens displayed by MHC
molecules
– TCRs have evolved to be specific for MHC molecules
– Majority of T cells recognize only peptides
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
8. ANTIGEN CAPTURE AND THE FUNCTIONS
OF ANTIGEN-PRESENTING CELLS
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
9. ANTIGEN CAPTURE AND THE FUNCTIONS OF
ANTIGEN-PRESENTING CELLS
• APC function is enhanced by exposure to microbial
products
– Toll-like receptors and other microbial sensors in dendritic
cells and macrophages
– Improved antigen presentation efficiency and APC cytokine
production Increase expression of MHC and
costimulators
– Adjuvants: products of microbes or mimic microbes
• Enhance expression of costimulators and cytokines
• Enhance functions of APC’s
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
10. ANTIGEN CAPTURE AND THE FUNCTIONS OF
ANTIGEN-PRESENTING CELLS
• APCs that present antigen to T cells also receive
signals from these Lymphocytes, enhancing their
antigen-presenting function
– Activated CD4+ express CD40L --- CD40 on dendritic
cells and macrophages IFN-γ secretion, activates
APC’s
• Leads to increased ability to process and present antigens,
• Increased expression of costimulators
• Secretion of cytokines
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
11. Role of Dendritic Cells in
Antigen Capture and Display
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
12. Role of Dendritic Cells in
Antigen Capture and Display
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
13. Role of Dendritic Cells in
Antigen Capture and Display
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
14. 2 Sets of Dendritic Cells
Classical DC • Most numerous subset of dendritic cells in the lymphoid
organs
• Mostly derived from myeloid precursors
• Constantly sample the environment
• May also present self antigens for regulation/self-tolerance.
• Upon encountering microbes/cytokines:
• Upregulate costimulatory molecules
• Produce inflammatory cytokines
• Migrate from peripheral tissue to draining lymph node
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
15. 2 Sets of Dendritic Cells
Classical DC • 2 subsets:
1. High expression of BDCA-1/CD1c – most potent at
driving CD4+ responses
2. Expression of BDCA-3 – efficient in process of cross-
presentation
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
16. 2 Sets of Dendritic Cells
Plasmacytoid
DC
• Resemble plasma cells
• Develop in Bone Marrow from same precursor as Classical
DC.
• Found in blood and in small numbers in lymphoid organs
• Poorly phagocytic and do NOT sample environmental
antigens
• Major function: Secretion of Type I IFN in response to
viral infections
• May also differentiate into cells similar to Classical DC and
present antigen to Virus-specific T-cells
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
17. Antigen Capture and Transport
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
Membrane Receptors
(C-type lectins)
Capture and Endocytose
microbes or microbial
products
Process ingested proteins
into peptides capable of
binding to MHC
18. Antigen Capture and Transport
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
Membrane Receptors
(C-type lectins)
Capture and Endocytose
microbes or microbial
products
Process ingested proteins
into peptides capable of
binding to MHC
Microbial products
recognized by TLR
Signals and Cytokines
activate DC
(TNF)
Activated DC lose
adhesiveness and migrate
to lymph nodes
19. Antigen Capture and Transport
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
DC
CCR7 Lymphatic Vessels
T cell zones of Lymph Nodes
CCL 19
CCL 21
Naïve
T cell
CCR7
“Colocalization”
20. Antigen Capture and Transport
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
DC
Capture Antigen
Present Antigen to Naïve
T cells
Activate Lymphocytes
Express high levels of
MHC
Activated DC develop
Into potent APCs
22. Antigen Capture and Transport
• Antigens can be transported to lymphoid
organs in soluble form
• Afferent Lymphatic Vessel Subcapsular
sinus FRC conduits Cortex
• Antigen can be extracted at the conduits,
some in the sinuses
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
23. Antigen Capture and Transport
Properties that make DC the most efficient APCs for
initiating T cell responses
1. Strategically located at common sites of entry
2. Express receptors that enable capture and response
3. Migrate from epithelia and tissues via lymphatics to T
cell zones of LN
4. Mature DC express high levels of peptide-MHC
complexes, costimulators, and cytokines
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
24. Antigen Capture and Transport
• Dendritic cells can also ingest infected cells
and present antigens to CD8+ T lymphocytes
– Peptide antigens must be derived from proteins in
the cytosol of DC
– Specialized DC: able to ingest virus-infected cells
and deliver viral proteins into their cytosol
– “Cross-presentation or Cross-priming”
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
25. Function of Other APCs
Cell-mediated Immune Responses Macrophages present Ag of phagocytosed
microbes to effector T cells
Humoral Immune Responses B lymphocytes internalize protein Ag and
present peptides from these proteins to helper T
cells.
Nucleated cells Can present peptides, derived from cytosolic
protein antigens CD8+ CTLs
Other cell types that express MHC
class II (endothelial and some
epithelial cells)
May present Ag to T cells
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
27. PROCESSING OF PROTEIN ANTIGENS
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
28. PROCESSING OF PROTEIN ANTIGENS
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
29. PROCESSING OF PROTEIN ANTIGENS
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
30. Class I MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
Injected via
Bacterial
secretory
mechanisms
Phagocytosed
Escape
Mechanism
31. Class I MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
32. Class I MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
33. Class I MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
34. Class I MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
Membrane
Chaperone:
Calnexin Luminal Chaperone:
Calreticulin
35. Class I MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
36. Class I MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
• Peptides transported into ER preferentially
bind to Class I MHC but NOT Class II MHC:
1. Class I attached to TAP complex
2. Class II molecules are blocked by a protein called
the invariant chain
37. Class I MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
38. Class I MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
40. Class II MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
Endosome-Lysosome
Phagolysosomes
Autophagosomes
41. Class II MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
CATHEPSINS
42. Class II MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
43. Class II MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
Membrane
Chaperone:
Calnexin
44. Class II MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
45. Class II MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
STEP 1:
Cathepsins degrade
Invariant Chain
CLIP STEP 2:
HLA-DM removes
CLIP
46. Class II MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
47. Class II MHC Pathway
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
53. Presentation of Non-Protein Antigens
• Small populations of T cells can recognize non-
protein antigens without Class I or II MHC:
NKT cells and γδ T cells.
– NKT: recognize lipids and glycolipids displayed by
CD1
– γδ T cells: recognize proteins, lipids,
phosphorylated molecules and alkyl amines
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
54. Outline
I. Properties of Antigens Recognized by T
Lymphocytes
II. Antigen Capture and the Functions of Antigen-
Presenting Cells
III. Processing of Protein Antigens
IV. Presentation of Non-protein Antigens to Subsets
of T Cells
55. Summary
1. Most T cells recognize antigens only as peptides displayed
by the products of self MHC genes on the surface of APCs.
2. MHC is a large genetic region coding for highly
polymorphic, co-dominantly expressed class I and class II
MHC molecules
3. The expression of MHC gene products is enhanced by
inflammatory and immune stimuli, particularly cytokines
like IFN-γ, which stimulate the transcription of MHC
genes.
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
56. Summary
MHC I MHC II
Composed of an α (or heavy) chain in a
non-covalent complex with a β2-
microglobulin
Contain two MHC-encoded polymorphic
chains, an α chain and a β chain.
Recognized by CD8+ T cells Recognized by CD4+ T cells
Accommodate peptides that are 6 to 16
amino acid residues in length
Allows larger peptides (up to 30 amino
acid residues in length or more) to bind
Expressed on all nucleated cells Expressed mainly on specialized APCs
Cytosolic proteins are proteolytically
degraded in the proteasome
Extracellular proteins are internalized into
endosomes
Cellular and Molecular Immunology 8th Ed. (2015) by Abbas et al.
58. References
• Cellular and Molecular
Immunology 8th Ed.
(2015) by Abbas et al.
– Chapter 6: MHC Molecules
and Antigen Presentation to T
Lymphocytes
Hinweis der Redaktion
The principal functions of T lymphocytes are to eradicate infections by intracellular microbes and to activate other cells, such as macrophages and B lymphocytes. To serve these functions, T cells have to overcome several challenges.
l There are very few naive T cells specific for any one antigen, and this small number has to be able to locate the foreign antigen and eliminate it.
Microbes and other antigens may be located at virtually any site in the body.
It is impossible for the few T cells specific for any antigen to constantly patrol all the possible tissues where antigens may enter or be produced.
Solving this problem requires a specialized system for capturing antigen and bringing it to the lymphoid organs through which T cells circulate and where responses can be initiated.
The specialized cells that capture and display antigens and activate T lymphocytes are called antigen- presenting cells (APCs).
l The functions of most T lymphocytes require that they interact with other cells, which may be dendritic cells, macrophages, B lymphocytes, or any infected host cell.
To ensure that T cells interact with other cells and not with soluble antigens, T cell antigen receptors are designed to see antigens displayed by cell surface molecules and not antigens on microbes or antigens that are free in the circulation or extracellular fluids.
This is in striking contrast to B lymphocytes, whose antigen receptors and secreted products, antibodies, can recognize antigens on microbial surfaces, and soluble antigens as well as cell-associated antigens.
The task of displaying host cell–associated antigens for recognition by CD4+ and CD8+ T cells is performed by specialized proteins called major histocompatibility complex (MHC) molecules, which are expressed on the surfaces of host cells.
l Different T cells have to be able to respond to microbial antigens in different cellular compartments.
For instance, defense against viruses in the circulation has to be mediated by antibodies, and the production of the most effective antibodies requires the participation of CD4+ helper T cells.
But if the same virus infects a tissue cell, it becomes inaccessible to the antibody, and its eradication requires that CD8+ cytotoxic T lymphocytes (CTLs) kill the infected cells and eliminate the reservoir of infection.
This dichotomy exists because APCs differentially handle antigens derived from extracellular or intracellular locations and present these antigens to the different classes of T cells.
MHC molecules play a critical role in segregating antigens from outside versus inside cells and displaying them to different T cell populations.
Thus, antigen capture and display to T cells is a specialized process that is essential for triggering optimal T cell responses.
Elucidation of the cell biology and molecular basis of this complex process has been an impressive accomplishment, based on functional experiments, biochemical analyses, and structural biology.
Most T lymphocytes recognize only short peptides, whereas B cells can recognize peptides, proteins, nucleic acids, carbohydrates, lipids, and small chemicals.
As a result, T cell–mediated immune responses are usually induced by foreign protein antigens (the natural source of foreign peptides),
whereas humoral immune responses are induced by protein and non-protein antigens.
The antigen receptors of CD4+ and CD8+ T cells are specific for peptide antigens that are displayed by MHC molecules (Fig. 6-1).
T cell receptors (TCRs) have evolved to be specific for MHC molecules, whose normal function is to display peptides.
MHC molecules can bind and display peptides and no other chemical structures, and this is why the majority of T cells recognize only peptides.
As we will dis- cuss later, MHC molecules are highly polymorphic, and variations in MHC molecules among individuals influence both peptide binding and T cell recognition.
A single T cell can recognize a specific peptide displayed by only one of the large number of different MHC molecules that exist.
This phenomenon is called MHC restriction, and we will describe its molecular basis later in this chapter.
TABLE 6-1
Different cell types function as APCs to activate naïve T cells or previously differentiated effector T cells.
Dendritic cells are the most effective APCs for activating naïve T cells
Macrophages and B lymphocytes – mostly for previously activated CD4+ helper T cells
These cells display MHC class II molecules
APCs display peptide-MHC complexes for recognition by T cells and provide additional stimuli required for full responses of the T cells
“second signals”/costimulators – additional stimuli for activation of naïve cells
APCs also secrete cytokines critical in T cell differentiation into effector cells
ANTIGEN CAPTURE AND THE FUNCTIONS OF ANTIGEN-PRESENTING CELLS
The antigen-presenting function of APCs is enhanced by exposure to microbial products.
This is one reason that the immune system responds better to microbes than to harmless, non-microbial substances.
Dendritic cells and macrophages express Toll-like receptors and other microbial sensors (see Chapter 4) that respond to microbes by increasing the expression of MHC molecules and costimulators, by improving the efficiency of antigen presentation, and by activating the APCs to produce cytokines, all of which stimulate T cell responses.
In addition, dendritic cells that are activated by microbes express chemokine receptors that stimulate their migration to sites where T cells are present.
The induction of optimal T cell responses to purified protein antigens requires that the antigens be administered with substances called adjuvants.
Adjuvants either are products of microbes, such as killed mycobacteria (used experimentally), or they mimic microbes and enhance the expression of costimulators and cytokines as well as the antigen-presenting functions of APCs.
ANTIGEN CAPTURE AND THE FUNCTIONS OF ANTIGEN-PRESENTING CELLS
APCs that present antigens to T cells also receive signals from these lymphocytes that enhance their antigen- presenting function.
In particular, CD4+ T cells that are activated by antigen recognition and costimulation express surface molecules, notably one called CD40 ligand (CD154), that binds to CD40 on dendritic cells and macrophages, and the T cells secrete cytokines, such as interferon-γ (IFN-γ), that bind to their receptors on these APCs.
The combination of CD40 signals and cytokines activates the APCs, resulting in increased ability to process and present antigens, increased expression of costimulators, and secretion of cytokines that activate the T cells.
This bidirectional interaction between APCs displaying the antigen and T lymphocytes that recognize the antigen functions as a positive feedback loop that plays an important role in maximizing the immune response (see Chapter 9).
Role of Dendritic Cells in Antigen Capture and Display
Common entry points for microbes are the skin, epithelia of the GI and respiratory organs
Role of Dendritic Cells in Antigen Capture and Display
Microbes and protein antigens are transported after being collected from their portal of entry into the peripheral lymphoid organs with the use of the numerous lymphatic capillaries that drain into the regional lymph nodes.
(Furthermore, if microbial antigens produced from any tissue colonized/infected by a microbe, )
Initiation of the primary responses of naïve T cells are done in these peripheral lymphoid organs
Some antigens are transported in the lymph by APCs (primarily dendritic cells) that capture the antigen and enter lymphatic vessels,
and other antigens enter the lymphatics in cell-free form.
Thus, the lymph contains a sampling of all the soluble and cell-associated antigens present in tissues.
The antigens become concentrated in lymph nodes,
Antigens that enter the blood stream may be similarly sampled by the spleen.
The cells that are best able to capture, transport, and present antigens to T cells are the dendritic cells.
Now lets discuss dendritic cells
l Classical DCs (also called conventional DCs) were first identified by their morphology and ability to stimulate strong T cell responses, and are the most numerous dendritic cell subset in lymphoid organs.
Most of them are derived from myeloid precursors, which migrate from the bone marrow to differentiate locally into resident dendritic cells in lymphoid and non- lymphoid tissues.
Similar to tissue macrophages, they constantly sample the environment in which they reside.
In the intestine, for example, dendritic cells appear to send out processes that traverse the epithelial cells and project into the lumen, where they may function to capture luminal antigens.
Langerhans cells are the dendritic cells that populate the epidermis; they serve the same role for antigens encountered in the skin.
In the absence of infection or inflammation, classical dendritic cells capture tissue antigens and migrate to the draining lymph nodes but do not produce cytokines and membrane molecules that
are required to induce effective immune responses.
The function of these dendritic cells may be to present self antigens to self-reactive T cells and thereby cause inactivation or death of the T cells or generate regulatory T cells.
These mechanisms are important for maintaining self- tolerance and preventing autoimmunity (see Chapter 15).
On encounter with microbes or cytokines, the dendritic cells become activated: they upregulate costimulatory molecules, produce inflammatory cytokines, and migrate from peripheral tissues into draining lymph nodes, where they initiate T cell responses (discussed later).
Classical dendritic cells may be divided into two major subsets.
One, identified by high expression of BDCA-1/ CD1c in humans or the CD11b integrin in mice, is most potent at driving CD4+ T cell responses.
The other sub- set, identified by expression of BDCA-3 in humans or, in mice, CD8 in lymphoid tissues or the CD103 integrin in peripheral tissues, is particularly efficient in the process of cross-presentation (described later in this chapter). Some dendritic cells may be derived from monocytes, especially in situations of inflammation.
Plasmacytoid DCs resemble plasma cells morphologically and acquire the morphology and functional proper- ties of dendritic cells only after activation.
They develop in the bone marrow from a precursor that also gives rise to classical dendritic cells, and are found in the blood and in small numbers in lymphoid organs.
In contrast to classical dendritic cells, plasmacytoid dendritic cells are poorly phagocytic and do not sample environmental antigens.
The major function of plasmacytoid dendritic cells is the secretion of large amounts of type I interferons in response to viral infections (see Chapter 4).
In viral infections, plasmacytoid dendritic cells also differentiate into cells that resemble classical dendritic cells and play a role in presenting antigens to virus-specific T cells.
Dendritic cells that are resident in epithelia and tissues capture protein antigens and transport the antigens to draining lymph nodes (Fig. 6-5).
Resting tissue-resident dendritic cells (sometimes referred to as immature dendritic cells) express membrane receptors, such as C-type lectins, that bind microbes.
Dendritic cells use these receptors to capture and endocytose microbes or microbial products and then process the ingested proteins into peptides capable of binding to MHC molecules.
Apart from receptor-mediated endocytosis and phagocytosis, dendritic cells can ingest antigens by micropinocytosis and macropinocytosis, processes that do NOT involve specific recognition receptors but capture whatever might be in the fluid phase in the vicinity of the dendritic cells.
Dendritic cells that are resident in epithelia and tissues capture protein antigens and transport the antigens to draining lymph nodes (Fig. 6-5).
At the time that microbial antigens are being captured, microbial products are recognized by Toll-like receptors and other innate pattern recognition receptors in the dendritic cells and other cells, generating innate immune responses
The dendritic cells are activated by these signals and by cytokines, such as tumor necrosis factor (TNF), produced in response to the microbes.
The activated dendritic cells (also called mature dendritic cells) lose their adhesiveness for epithelia or tissues and migrate into lymph nodes.
The dendritic cells also begin to express a chemokine receptor called CCR7 that is specific for two chemokines, CCL19 and CCL21, which are produced in lymphatic vessels and in the T cell zones of lymph nodes.
These chemokines attract the dendritic cells bearing microbial antigens into draining lymphatics and ultimately into the T cell zones of the regional lymph nodes.
Naive T cells also express CCR7, and this is why naive T cells migrate to the same regions of lymph nodes where antigen-bearing dendritic cells are concentrated (see Chapter 3).
The colocalization of antigen-bearing activated dendritic cells and naive T cells maximizes the chance of T cells with receptors for the antigen finding that antigen.
Activation also converts the dendritic cells from cells whose primary function is to capture antigen into cells that are able to present antigens to naive T cells and to activate the lymphocytes.
Activated dendritic cells express high levels of MHC molecules with bound peptides as well as costimulators required for T cell activation.
Thus, by the time these cells arrive in the lymph nodes, they have developed into potent APCs with the ability to activate T lymphocytes.
Naive T cells that recirculate through lymph nodes encounter these APCs, and the T cells that are specific for the displayed peptide-MHC complexes are activated.
This is the initial step in the induction of T cell responses to protein antigens.
Antigen Capture and Transport by Dendritic Cells
Antigens may also be transported to lymphoid organs in soluble form. Resident dendritic cells in the lymph nodes and spleen may capture lymph- and blood-borne antigens, respectively, and also may be driven to mature by microbial products.
When lymph enters a lymph node through an afferent lymphatic vessel, it drains into the subcapsular sinus, and some of the lymph enters fibroblast reticular cell (FRC) conduits that originate from the sinus and traverse the cortex (see Chapter 2).
Once in the conduits, low–molecular-weight antigens can be extracted by dendritic cells whose processes interdigitate between the FRCs.
Other antigens in the subcapsular sinus are taken up by macrophages and dendritic cells, which carry the antigens into the cortex.
B cells in the node may also recognize and internalize soluble antigens.
Dendritic cells, macrophages, and B cells that have taken up protein antigens can then process and present these antigens to naive T cells and to effector T cells that have been generated by previous antigen stimulation.
The collection and concentration of foreign antigens in lymph nodes are supplemented by other anatomic adaptations that serve similar functions.
The mucosal surfaces of the gastrointestinal and respiratory systems, in addition to being drained by lymphatic capillaries, contain specialized collections of secondary lymphoid tissue that can directly sample the luminal contents of these organs for the presence of antigenic material.
The best characterized of these mucosal lymphoid organs are Peyer’s patches of the ileum and the pharyngeal tonsils (see Chapter 14).
APCs in the spleen monitor the blood stream for any antigens that reach the circulation.
Such antigens may reach the blood either directly from the tissues or by way of the lymph from the thoracic duct.
Antigen-Presenting Function of Dendritic Cells
Many studies done in vitro and in vivo have established that the induction of primary T cell–dependent immune responses to protein antigens requires the presence of dendritic cells to capture and to present the antigens to the T cells. This was first shown for CD4+ T cell responses but is now known to be true for CD8+ T cells as well.
Several properties of dendritic cells make them the most efficient APCs for initiating primary T cell responses.
l Dendritic cells are strategically located at the common sites of entry of microbes and foreign antigens (in epithelia) and in tissues that may be colonized by microbes.
l Dendritic cells express receptors that enable them to capture and respond to microbes.
l Dendritic cells migrate from epithelia and tissues via lymphatics preferentially into the T cell zones of lymph nodes, and naive T lymphocytes also migrate from the circulation into the same regions of the lymph nodes.
l Mature dendritic cells express high levels of peptide- MHC complexes, costimulators, and cytokines, all of which are needed to activate naive T lymphocytes.
Dendritic cells can ingest infected cells and present antigens from these cells to CD8+ T lymphocytes.
Dendritic cells are the best APCs to induce the primary responses of CD8+ T cells, but this poses a special problem because the peptide antigens these lymphocytes recognize must be derived from proteins in the cytosol of the dendritic cells.
However, the viral proteins may be produced in any cell type infected by a virus, not necessarily in dendritic cells.
Some specialized dendritic cells have the ability to ingest virus-infected cells or cellular fragments, and deliver viral proteins into their cytosol, allowing them to be presented to CD8+ T lymphocytes.
This process is called cross-presentation, or cross-priming, and is described later in this chapter.
Functions of Other Antigen-Presenting Cells
Although dendritic cells have a critical role in initiating primary T cell responses, other cell types are also important APCs in different situations (see Fig. 6-2 and Table 6-2).
l In cell-mediated immune responses, macrophages present the antigens of phagocytosed microbes to effector T cells, which respond by activating the macrophages to kill the microbes.
This process is central to cell- mediated immunity and delayed-type hypersensitivity (see Chapter 10).
Circulating monocytes are able to migrate to any site of infection and inflammation, where they differentiate into macrophages and phagocytose and destroy microbes.
CD4+ T cells recognize microbial antigens being presented by the macrophages and provide signals that enhance the microbicidal activities of these macrophages.
l In humoral immune responses, B lymphocytes internalize protein antigens and present peptides derived from these proteins to helper T cells. This antigen- presenting function of B cells is essential for helper T cell–dependent antibody production (see Chapter 12).
l All nucleated cells can present peptides, derived from cytosolic protein antigens, to CD8+ CTLs.
All nucleated cells are susceptible to viral infections and cancer-causing mutations.
Therefore, it is important that the immune system be able to recognize cytosolic antigens, such as viral antigens and mutated proteins, in any cell type.
CD8+ CTLs are the cell population that recognizes these antigens and eliminates the cells in which the antigens are produced.
CD8+ CTLs may also recognize phagocytosed microbes if these microbes or their antigens escape from phagocytic vesicles into the cytosol.
l Other cell types that express class II MHC molecules and may present antigens to T cells include endothelial and some epithelial cells.
Vascular endothelial cells may present antigens to blood T cells that adhere to vessel walls, and this process may contribute to the recruitment and activation of effector T cells in cell-mediated immune reactions.
Endothelial cells in grafts are also targets of T cells reacting against graft antigens (see Chapter 17).
Various epithelial and mesenchymal cells may express class II MHC molecules in response to the cytokine IFN-γ. The physiologic significance of antigen presentation by these cell populations is unclear.
Because most of them do not express costimulators and are not efficient at processing proteins into MHC- binding peptides, it is unlikely that they contribute significantly to the majority of T cell responses.
Thymic epithelial cells constitutively express MHC molecules and play a critical role in presenting peptide-MHC complexes to maturing T cells in the thymus as part of the selection processes that shape the repertoire of T cell specificities (see Chapter 8).
PROCESSING OF PROTEIN ANTIGENS
The pathways of antigen processing convert protein antigens present in the cytosol or internalized from the extracellular environment into peptides and load these peptides onto MHC molecules for display to T lymphocytes (Fig. 6-14).
The mechanisms of antigen processing are designed to generate peptides that have the structural characteristics required for associating with MHC molecules, and to place these peptides in the same cellular location as newly formed MHC molecules with available peptide-binding clefts.
Peptide binding to MHC molecules occurs before cell surface expression and is an integral component of the biosynthesis and assembly of MHC molecules.
In fact, as mentioned earlier, peptide association is required for the stable assembly and surface expression of class I and class II MHC molecules.
Protein antigens that are present in the cytosol (usually synthesized in the cell) generate class I–associated peptides that are recognized by CD8+ T cells,
whereas antigens internalized from the extracellular environment into the vesicles of APCs usually generate peptides that are displayed by class II MHC molecules and recognized by CD4+ T cells.
The different fates of cytosolic and vesicular antigens are due to the segregated pathways of biosynthesis and assembly of class I and class II MHC molecules (see Fig. 6-14 and Table 6-5).
The fundamental difference between the class I MHC and class II MHC pathways resides mainly in the site of peptide degradation.
Proteins degraded in proteasomes, many of which are from the cytosol, primarily provide peptides for class I MHC molecules.
Only proteins that are degraded in endo-lysosomes provide peptides for class II MHC molecules.
The difference between cytosolic and vesicular antigens has been demonstrated experimentally by analyzing the presentation of the same antigen introduced into APCs in different ways (Fig. 6-15).
If a protein antigen is produced in the cytoplasm of APCs as the product of a transfected gene (modified so its protein product cannot enter the secretory pathway) or introduced directly into the cytoplasm of the APCs by osmotic shock, peptides derived from the protein are presented by class I MHC molecules and are recognized by CD8+ T cells.
In contrast, if the same protein is added in soluble form to APCs and endocytosed into the vesicles of the APCs, peptides (which may be different from the ones presented by class I) are subsequently presented by class II molecules and are recognized by antigen-specific CD4+ T cells.
The Class I MHC Pathway for Processing and Presentation of Cytosolic Proteins
Class I MHC–associated peptides are produced by the proteolytic degradation of mainly cytosolic proteins in proteasomes, and the generated peptides are transported into the endoplasmic reticulum (ER), where they bind to newly synthesized class I molecules.
Sources of Cytosolic Protein Antigens
Most cytosolic protein antigens are synthesized within cells, some are injected into the cytosol via bacterial secretory mechanisms, and others are phagocytosed and transported from vesicles into the cytosol.
Foreign antigens in the cytosol may be the products of viruses, bacteria, or other intracellular microbes that infect such cells.
In tumor cells, various mutated or overexpressed genes may produce protein antigens that are recognized by class I–restricted CTLs (see Chapter 18).
Peptides that are presented in association with class I molecules may also be derived from microbes and other particulate antigens that are internalized into phagosomes but escape into the cytosol.
Some microbes are able to damage phagosome membranes and create pores through which the microbes and their antigens enter the cytosol.
For instance, pathogenic strains of Listeria monocytogenes produce a protein called listeriolysin that enables bacteria to escape from vesicles into the cytosol.
(This escape is a mechanism that the bacteria may have evolved to resist killing by the microbicidal mechanisms of phagocytes, most of which are concentrated in phagolysosomes.)
Once the antigens of the phagocytosed microbes are in the cytosol, they are processed like other cytosolic antigens.
AS LONG AS ANTIGEN IS IN THE CYTOSOL
In dendritic cells, some antigens that are ingested into vesicles enter the cytosolic class I pathway, in the process called cross-presentation that is described later.
Although microbial proteins that are presented on class I MHC molecules are typically cytosolic, proteins from other cellular compartments may also enter the class I MHC antigen processing pathway.
The signal sequences of membrane and secreted proteins are usually cleaved by signal peptidase and degraded proteolytically soon after synthesis and translocation into the ER.
This ER processing generates class I–binding peptides without a need for proteolysis in the cytosol.
In addition, nuclear proteins may be processed by proteasomes in the nucleus and presented on class I MHC molecules.
Digestion of Proteins in Proteasomes
The major mechanism for the generation of peptides from cytosolic and nuclear protein antigens is proteolysis by the proteasome.
Proteasomes are large multiprotein enzyme complexes with a broad range of proteolytic activity that are found in the cytoplasm and nuclei of most cells.
The proteasome appears as a cylinder composed of a stacked array of two inner β rings and two outer α rings, each ring being composed of seven subunits, with a cap-like structure at either end of the cylinder.
The proteins in the outer α rings are structural and lack proteolytic activity;
in the inner β rings, three of the seven subunits (β1, β2, and β5) are the catalytic sites for proteolysis.
The proteasome performs a basic housekeeping function in cells by degrading many damaged or improperly folded proteins.
Protein synthesis normally occurs at a rapid rate, about six to eight amino acid residues being incorporated into elongating chains every second.
The process is error prone, and it is estimated that approximately 20% of newly synthesized proteins are misfolded.
These newly translated but defective polypeptides, as well as proteins that are damaged by cellular stresses, are targeted for proteasomal degradation by covalent linkage of several copies of a small polypeptide called ubiquitin.
Digestion of Proteins in Proteasomes
Ubiquitinated proteins, with chains of four or more ubiquitins, are recognized by the proteasomal cap and then are unfolded, the ubiquitin is removed, and the proteins are threaded through proteasomes, where they are degraded into peptides.
The proteasome has broad substrate specificity and can generate a wide variety of peptides from cytosolic proteins (but usually does not degrade them completely into single amino acids).
There are mechanisms wherein IFN-γ can ENHANCE antigen presentation
By increased expression of MHC molecules
Increased transcription and synthesis of 3 novel catalytic subunits of proteosome:
known as β1i, β2i, and β5i, which replace the three catalytic subunits of the β ring of the proteasome.
This results in a change in the substrate specificity of the proteasome such that the peptides produced usually contain carboxy-terminal hydrophobic amino acids such as leucine, valine, isoleucine, and methionine or basic residues such as lysine or arginine.
These kinds of C termini are typical of peptides that are transported into the class I pathway and bind to class I molecules.
Thus, proteasomes are organelles whose basic cellular function has been adapted for a specialized role in antigen presentation.
Transport of Peptides from the Cytosol to the Endoplasmic Reticulum
Peptides generated in proteasomes are translocated by a specialized transporter into the ER, where newly synthesized class I MHC molecules are available to bind the peptides.
This delivery is mediated by a dimeric protein called transporter associated with antigen processing (TAP), which is a member of the ABC transporter family of proteins, many of which mediate ATP-dependent transport of low–molecular-weight compounds across cellular membranes.
The TAP protein is located in the ER membrane, where it mediates the active, ATP-dependent transport of peptides from the cytosol into the ER lumen.
Although the TAP heterodimer has a broad range of specificities, it optimally transports peptides ranging from 8 to 16 amino acids in length and containing carboxyl termini that are basic (in humans) or hydrophobic (in humans and mice).
As mentioned earlier, these are the characteristics of the peptides that are generated in the proteasome and are able to bind to class I MHC molecules.
On the luminal side of the ER membrane, the TAP protein associates with a protein called tapasin, which also has an affinity for newly synthesized empty class I MHC molecules.
Tapasin thus brings the TAP transporter into a complex with the class I MHC molecules that are awaiting the arrival of peptides.
Assembly of Peptide–Class I MHC Complexes in the Endoplasmic Reticulum
Peptides translocated into the ER bind to class I MHC molecules that are associated with the TAP dimer through tapasin.
The synthesis and assembly of class I molecules involve a multistep process in which peptide binding plays a key role.
Class I α chains and β2-microglobulin are synthesized in the ER.
Appropriate folding of the nascent α chains is assisted by chaperone proteins, such as the membrane chaperone calnexin and the luminal chaperone calreticulin.
Assembly of Peptide–Class I MHC Complexes in the Endoplasmic Reticulum
Within the ER, the newly formed empty class I dimers remain linked to the TAP complex.
Empty class I MHC molecules, tapasin, and TAP are part of a larger peptide-loading complex in the ER that also includes calnexin, calreticulin, and other components that contribute to class I MHC assembly and loading.
Peptides that enter the ER through TAP and peptides produced in the ER, such as signal peptides, are often trimmed to the appropriate size for MHC binding by the ER-resident amino- peptidase (ERAP).
The peptide is then able to bind to the cleft of the adjacent class I molecule.
Once class I MHC molecules are loaded with peptide, they no longer have an affinity for tapasin, so the peptide–class I complex is released, and it is able to exit the ER and be transported to the cell surface.
In the absence of bound peptide, many of the newly formed α chain–β2-microglobulin dimers are unstable and cannot be transported efficiently from the ER to the Golgi complex.
These misfolded empty class I MHC complexes are transported into the cytosol and degraded in the proteasomes.
Peptides transported into the ER preferentially bind to class I but not class II MHC molecules for two reasons.
First, newly synthesized class I molecules are attached to the luminal aspect of the TAP complex, and they capture peptides rapidly as the peptides are transported into the ER by the TAP.
Second, as discussed later, in the ER the peptide-binding clefts of newly synthesized class II molecules are blocked by a protein called the invariant chain.
Surface Expression of Peptide–Class I MHC Complexes
Class I MHC molecules with bound peptides are structurally stable and are expressed on the cell surface.
Stable peptide–class I MHC complexes that were produced in the ER move through the Golgi complex and are transported to the cell surface by exocytic vesicles.
Once expressed on the cell surface, the peptide–class I complexes may be recognized by peptide antigen–specific CD8+ T cells, with the CD8 coreceptor playing an essential role by binding to non-polymorphic regions of the class I molecule.
SUMMARIZE
The Class II MHC Pathway for Processing and Presentation of Vesicular Proteins
The generation of class II MHC–associated peptides from endocytosed antigens involves the proteolytic degradation of internalized proteins in endocytic vesicles and the binding of peptides to class II MHC molecules in vesicles.
Generation of Vesicular Proteins
Most class II MHC–associated peptides are derived from protein antigens that are captured from the extracellular environment and internalized into endosomes by specialized APCs.
The initial steps in the presentation of an extracellular protein antigen are the binding of the native antigen to an APC and the internalization of the antigen.
Different APCs can bind protein antigens in several ways and with varying efficiencies and specificities.
Dendritic cells and macrophages express a variety of surface receptors that recognize structures shared by many microbes (see Chapter 4).
These APCs use the receptors to bind and internalize microbes efficiently.
Macrophages also express receptors for the Fc portions of antibodies and receptors for the complement protein C3b, which bind antigens with attached antibodies or complement proteins and enhance their internalization.
Another example of specific receptors on APCs is the surface immunoglobulin on B cells, which, because of its high affinity for antigens, can effectively mediate the internalization of proteins present at very low concentrations in the extracellular fluid (see Chapter 12).
After their internalization, protein antigens become localized in intracellular membrane-bound vesicles called endosomes.
The endosomal pathway of intracellular protein traffic communicates with lysosomes, which are denser membrane-bound enzyme-containing vesicles.
A subset of class II MHC–rich late endosomes plays a special role in antigen processing and presentation by the class II pathway; this is described later.
Particulate microbes are internalized into vesicles called phagosomes, which may fuse with lysosomes, producing vesicles called phagolysosomes or secondary lysosomes.
Proteins other than those ingested from the extracellular milieu can also enter the class II MHC pathway.
Some protein molecules destined for secretion may end up in the same vesicles as class II MHC molecules and may be processed instead of being secreted.
Less often, cytoplasmic and membrane proteins may be processed and displayed by class II molecules.
In some cases, this may result from the enzymatic digestion of cytoplasmic contents, referred to as autophagy.
In this pathway, cytosolic proteins are trapped within membrane-bound vesicles called autophagosomes; these vesicles fuse with lysosomes, and the cytoplasmic proteins are proteolytically degraded.
The peptides generated by this route may be delivered to the same class II–bearing vesicular compartment as are peptides derived from ingested antigens.
Autophagy is primarily a mechanism for degrading cellular proteins and recycling their products as sources of nutrients during times of stress.
It also participates in the destruction of intracellular microbes, which are enclosed in vesicles and delivered to lysosomes.
It is therefore predictable that peptides generated by autophagy will be displayed for T cell recognition.
Some peptides that associate with class II molecules are derived from membrane proteins, which may be recycled into the same endocytic pathway as are extracellular proteins.
Thus, even viruses, which replicate in the cytoplasm of infected cells, may produce proteins that are degraded into peptides that enter the class II MHC pathway of antigen presentation.
This may be a mechanism for the activation of viral antigen–specific CD4+ helper T cells.
Proteolytic Digestion of Proteins in Vesicles
Internalized proteins are degraded enzymatically in late endosomes and lysosomes to generate peptides that are able to bind to the peptide-binding clefts of class II MHC molecules.
The degradation of protein antigens in vesicles is an active process mediated by proteases that have acidic pH optima.
The most abundant proteases of late endosomes are cathepsins, which are thiol and aspartyl proteases with broad substrate specificities.
Several cathepsins contribute to the generation of peptides for the class II pathway.
Partially degraded or cleaved proteins bind to the open-ended clefts of class II MHC molecules and are then trimmed enzymatically to their final size.
Proteolytic Digestion of Proteins in Vesicles
studies have defined a class II MHC–rich subset of late endosomes that plays an important role in antigen presentation (Fig. 6-18).
In macrophages and human B cells, it is called the MHC class II compartment, or MIIC.
The MIIC has a characteristic multilamellar appearance by electron microscopy.
Importantly, it contains all of the components required for peptide–class II MHC association, including the
enzymes that degrade protein antigens,
class II MHC molecules,
and two molecules involved in peptide loading of class II MHC molecules,
the invariant chain and HLA-DM,
whose functions are described later.
Biosynthesis and Transport of Class II MHC Molecules to Endosomes
Class II MHC molecules are synthesized in the ER and transported to endosomes with an associated protein, the invariant chain, which occupies the peptide-binding clefts of the newly synthesized class II MHC molecules (Fig. 6-19).
The α and β chains of class II MHC molecules are coordinately synthesized and associate with each other in the ER.
Nascent class II MHC dimers are structurally unstable, and their folding and assembly are aided by ER- resident chaperones, such as calnexin.
Biosynthesis and Transport of Class II MHC Molecules to Endosomes
The invariant chain (Ii) promotes folding and assembly of class II MHC molecules and directs newly formed class II MHC molecules to the late endosomes and lysosomes where internalized proteins have been proteolytically degraded into peptides.
The invariant chain is a trimer composed of three 30-kD subunits, each of which binds one newly synthesized class II MHC αβ heterodimer in a way that blocks the peptide-binding cleft and prevents it from accepting peptides.
As a result, class II MHC molecules cannot bind and present peptides they encounter in the ER, leaving such peptides to associate with class I molecules (described earlier).
The class II MHC molecules are transported in exocytic vesicles toward the cell surface.
During this passage, the vesicles taking class II MHC molecules out of the ER meet and fuse with the endocytic vesicles containing internalized and processed antigens.
Thus, class II MHC molecules encounter antigenic peptides that have been generated by proteolysis of endocytosed proteins, and the peptide-MHC association occurs in the vesicles.
Association of Processed Peptides with Class II MHC Molecules in Vesicles
Within the endosomal vesicles, the invariant chain dissociates from class II MHC molecules by the combined action of proteolytic enzymes and the HLA-DM molecule, and antigenic peptides are then able to bind to the available peptide-binding clefts of the class II molecules (see Fig. 6-19).
Because the invariant chain blocks access to the peptide-binding cleft of class II MHC molecules, it must be removed before complexes of peptide and class II MHC molecules can form.
The same proteolytic enzymes that generate peptides from internalized proteins, such as cathepsins, also act on the invariant chain, degrading it and leaving only a 24–amino acid remnant called class II–associated invariant chain peptide (CLIP), which sits in the peptide-binding cleft in the same way that other peptides bind to class II MHC molecules.
Next, CLIP must be removed so that the cleft becomes accessible to antigenic peptides produced from extracellular proteins.
This removal is accomplished by the action of a molecule called HLA-DM (or H-2M in the mouse), which is encoded within the MHC, has a structure similar to that of class II MHC molecules, and colocalizes with class II MHC molecules in the MIIC endosomal compartment.
Unlike class II MHC molecules, HLA-DM molecules are not polymorphic, and they are not expressed on the cell surface.
HLA- DM acts as a peptide exchanger, facilitating the removal of CLIP and the addition of other peptides to class II MHC molecules.
If peptides with a higher affinity for the class II MHC cleft than CLIP are available in endosomes they will be able to displace CLIP because of the HLA-DM mediated exchange mechanism.
If higher affinity peptides are not available CLIP will remain in the class II MHC cleft and these molecules will not undergo the presumed conformational change and stabilization required to efficiently travel to the cell surface.
Because the ends of the class II MHC peptide-binding cleft are open, large peptides may bind and are then trimmed by proteolytic enzymes to the appropriate size for T cell recognition.
As a result, the peptides that are actually presented attached to cell surface class II MHC molecules are usually 10 to 30 amino acids long and typically have been generated by this trimming step.
Expression of Peptide–Class II MHC Complexes on the Cell Surface
Class II MHC molecules are stabilized by the bound peptides, and the stable peptide–class II complexes are delivered to the surface of the APC, where they are displayed for recognition by CD4+ T cells.
The transport of class II MHC–peptide complexes to the cell surface is believed to occur by fusion of vesiculotubular extensions from the lysosome with the plasma membrane, resulting in delivery of the loaded class II MHC complexes to the cell surface.
Once expressed on the APC surface, the peptide–class II complexes are recognized by peptide antigen–specific CD4+ T cells, with the CD4 coreceptor playing an essential role by binding to non-polymorphic regions of the class II molecule.
Interestingly, whereas peptide-loaded class II molecules traffic from the late endosomes and lysosomes to the cell surface, other molecules involved in antigen presentation, such as DM, stay in the vesicles and are not expressed in the plasma membrane.
The mechanism of this selective traffic is unknown.
SUMMARIZE
Cross-Presentation
Some dendritic cells have the ability to capture and to ingest virus-infected cells or tumor cells and present the viral or tumor antigens to naive CD8+ T lymphocytes Fig. 6-20).
In this pathway, the ingested antigens are transported from vesicles to the cytosol, from where peptides enter the class I pathway.
As we discussed earlier, most ingested proteins do not enter the cytosolic class I pathway of antigen presentation.
This permissiveness for protein traffic from endosomal vesicles to the cytosol is unique to dendritic cells.
(At the same time, the dendritic cells can present class II MHC–associated peptides generated in the vesicles to CD4+ helper T cells, which are often required to induce full responses of CD8+ cells [see Chapter 11].)
This process is called cross-presentation, or cross-priming, to indicate that one cell type (the dendritic cell) can present antigens from another cell (the virus-infected or tumor cell) and prime, or activate, T cells specific for these antigens.
Although on face value it may seem that the process of cross-presentation violates the rule that ingested antigens are presented bound to class II MHC molecules, in this situation the ingested antigens are degraded in proteasomes and enter the class I pathway.
Cross-presentation involves the fusion of phagosomes containing the ingested antigens with the ER.
Ingested proteins are then translocated from the ER to the cytosol by poorly defined pathways that are likely involved in presentation of proteins degraded in the ER.
The proteins that were initially internalized in the phagosome are therefore delivered to the compartment (the cytosol) where proteolysis for the class I pathway normally occurs.
These phagocytosed proteins thus undergo proteasomal degradation, and peptides derived from them are transported by TAP back into the ER, where they are assembled with newly synthesized class I MHC molecules as described for the conventional class I pathway.
Nature of T Cell Responses
The presentation of cytosolic versus vesicular proteins by the class I or class II MHC pathways, respectively, determines which subsets of T cells will respond to antigens found in these two pools of proteins and is intimately linked to the functions of these T cells (Fig. 6-21).
Endogenously synthesized antigens, such as viral and tumor proteins, are located in the cytosol and are recognized by class I MHC–restricted CD8+ CTLs, which kill the cells producing the intracellular antigens.
Conversely, extracellular antigens usually end up in endosomal vesicles and activate class II MHC–restricted CD4+ T cells because vesicular proteins are processed into class II–binding peptides.
CD4+ T cells function as helpers to stimulate B cells to produce antibodies and activate macrophages to enhance their phagocytic functions, both mechanisms that serve to eliminate extracellular antigens.
Thus, antigens from microbes that reside in different cellular locations selectively stimulate the T cell responses that are most effective at eliminating that type of microbe.
This is especially important because the antigen receptors of CTLs and helper T cells cannot distinguish between extracellular and intracellular microbes.
By segregating peptides derived from these types of microbes, the MHC molecules guide CD4+ and CD8+ subsets of T cells to respond to the microbes that each subset can best combat.
Immunogenicity of Protein Antigens
MHC molecules determine the immunogenicity of protein antigens in two related ways.
The epitopes of complex proteins that elicit the strongest T cell responses are the peptides that are generated by proteolysis in APCs and bind most avidly to MHC molecules.
If an individual is immunized with a protein antigen, in many instances the majority of the responding T cells are specific for only one or a few linear amino acid sequences of the antigen.
These are called the immunodominant epitopes or determinants.
The proteases involved in antigen processing produce a variety of peptides from natural proteins, and only some of these peptides possess the characteristics that enable them to bind to the MHC molecules present in each individual (Fig. 6-22).
It is important to define the structural basis of immunodominance because this may permit the efficient manipulation of the immune system with synthetic peptides.
An application of such knowledge is the design of vaccines.
For example, a viral protein could be analyzed for the presence of amino acid sequences that would form typical immunodominant epitopes capable of binding to MHC molecules with high affinity.
Synthetic peptides containing these epitopes may be effective vaccines for eliciting T cell responses against the viral peptide expressed on an infected cell.
The expression of particular class II MHC alleles in an individual determines the ability of that individual to respond to particular antigens.
As discussed earlier, the immune response (Ir) genes that control antibody responses are the class II MHC genes.
They influence immune responsiveness because various allelic class II MHC molecules differ in their ability to bind different antigenic peptides and therefore to stimulate specific helper T cells.
The consequences of inheriting a given MHC allele depend on the nature of the peptide antigens that can bind the MHC molecule encoded by that allele.
For example, if the antigen is a peptide from ragweed pollen, the individual who expresses class II molecules capable of binding the peptide would be genetically prone to allergic reactions against pollen.
Conversely, some individuals do not respond to vaccines (such as hepatitis B virus surface antigen vaccine), presumably because their HLA molecules cannot bind and display the major peptides of the antigen.
PRESENTATION OF NON-PROTEIN ANTIGENS TO SUBSETS OF T CELLS
Several small populations of T cells are able to recognize non-protein antigens without the involvement of class I or class II MHC molecules.
Thus, these populations are exceptions to the rule that T cells can see only MHC- associated peptides.
The best defined of these populations are NKT cells and γδ T cells.
NKT cells express markers that are characteristic of both natural killer (NK) cells and T lymphocytes and express αβ T cell receptors with very limited diversity (see Chapter 10).
NKT cells recognize lipids and glycolipids displayed by the class I–like non-classical MHC molecule called CD1.
There are several CD1 proteins expressed in humans and mice.
Although their intracellular traffic pathways differ in subtle ways, all CD1 molecules bind and display lipids by a unique mechanism.
Newly synthesized CD1 molecules pick up cellular lipids and carry these to the cell surface.
From here, the CD1-lipid complexes are endocytosed into endosomes or lysosomes, where lipids that have been ingested from the external environment are captured and the new CD1-lipid complexes are returned to the cell surface.
Thus, CD1 molecules acquire endocytosed lipid antigens during recycling and present these antigens without apparent processing.
The NKT cells that recognize the lipid antigens may play a role in defense against microbes, especially mycobacteria (which are rich in lipid components).
γδ T cells are a small population of T cells that express antigen receptor proteins that are similar but not identical to those of CD4+ and CD8+ T cells (see Chapter 10).
γδ T cells recognize many different types of antigens, including some proteins and lipids, as well as small phosphorylated molecules and alkyl amines.
These antigens are not displayed by MHC molecules, and γδ cells are not MHC restricted.
It is not known if a particular cell type or antigen display system is required for presenting antigens to these cells.
Most T cells recognize antigens only in the form of peptides displayed by the products of self MHC genes on the surface of APCs. CD4+ helper T lymphocytes recognize antigens in association with class II MHC gene products (class II MHC–restricted recognition), and CD8+ CTLs recognize anti- gens in association with class I MHC gene products (class I MHC–restricted recognition).
Specialized APCs, such as dendritic cells, macrophages, and B lymphocytes, capture extracellular protein antigens, internalize and process them, and display class II–associated peptides to CD4+ T cells. Dendritic cells are the most efficient APCs for initiating primary responses by activating naive T cells, and macrophages and B lymphocytes present antigens to differentiated helper T cells in the effector phase of cell-mediated immunity and in humoral immune responses, respectively. All nucleated cells can present class I–associated peptides, derived from cytosolic proteins such as viral and tumor antigens, to CD8+ T cells.
The MHC is a large genetic region coding for highly polymorphic, codominantly expressed class I and class II MHC molecules.
Class I MHC molecules are composed of an α (or heavy) chain in a non-covalent complex with a non-polymorphic polypeptide called β2- microglobulin. Class II MHC molecules contain two MHC-encoded polymorphic chains, an α chain and a β chain. Both classes of MHC molecules consist of an extracellular peptide-binding cleft, a non-polymorphic Ig-like region, a transmembrane region, and a cytoplasmic region. The peptide-binding cleft of MHC molecules has α-helical sides and an eight-stranded antiparallel β-pleated sheet floor. The Ig-like domains of class I and class II MHC molecules contain the binding sites for the T cell coreceptors CD8 and CD4, respectively. The polymorphic residues of MHC molecules are localized to the peptide-binding domain.
The function of class I and class II MHC molecules is to bind peptide antigens and display them for recognition by antigen-specific T lymphocytes. Peptide antigens associated with class I MHC molecules are recognized by CD8+ T cells, whereas class II MHC– associated peptide antigens are recognized by CD4+ T cells. MHC molecules bind only one peptide at a time, and all of the peptides that bind to a particular MHC molecule share common structural motifs. Every MHC molecule has a broad specificity for peptides and can bind multiple peptides that have common structural features, such as anchor residues.
The peptide-binding cleft of class I MHC molecules can accommodate peptides that are 6 to 16 amino acid residues in length, whereas the cleft of class II MHC molecules allows larger peptides (up to 30 amino acid residues in length or more) to bind. Some polymorphic MHC residues determine the binding specificities for peptides by forming structures called pockets that interact with complementary residues of the bound pep- tide, called anchor residues. Other polymorphic MHC residues and some residues of the peptide are not involved in binding to MHC molecules but instead form the structure recognized by T cells.
Class I MHC molecules are expressed on all nucleated cells, whereas class II MHC molecules are expressed mainly on specialized APCs, such as dendritic cells, macrophages, and B lymphocytes, and a few other cell types, including endothelial cells and thymic epithelial cells. The expression of MHC gene products is enhanced by inflammatory and immune stimuli, particularly cytokines like IFN-γ, which stimulate the transcription of MHC genes.
Antigen processing is the conversion of native proteins into MHC-associated peptides. This process consists of the introduction of exogenous protein antigens into vesicles of APCs or the synthesis of antigens in the cytosol, the proteolytic degradation of these proteins into peptides, the binding of peptides to MHC molecules, and the display of the peptide-MHC complexes on the APC surface for recognition by T cells. Thus, both extracellular and intracellular proteins are sampled by these antigen-processing pathways, and peptides derived from both normal self proteins and foreign proteins are displayed by MHC molecules for surveillance by T lymphocytes.
For the class I MHC pathway, cytosolic proteins are proteolytically degraded in the proteasome, generating peptides that bind to class I MHC molecules. These peptides are delivered from the cytosol to the ER by an ATP-dependent transporter called TAP. Newly synthesized class I MHC–β2- microglobulin dimers in the ER are associated with the TAP complex and receive peptides trans- ported into the ER. Stable complexes of class I MHC molecules with bound peptides move out of the ER, through the Golgi complex, to the cell surface.
For the class II MHC pathway, extracellular proteins are internalized into endosomes, where these proteins are proteolytically cleaved by enzymes that function at acidic pH. Newly synthesized class II MHC molecules associated with the Ii are trans- ported from the ER to the endosomal vesicles. Here the Ii is proteolytically cleaved, and a small peptide remnant of the Ii, called CLIP, is removed from the peptide-binding cleft of the MHC molecule by the DM molecules. The peptides that were generated from extracellular proteins then bind to the available cleft of the class II MHC molecule, and the trimeric complex (class II MHC α and β chains and peptide) moves to and is displayed on the surface of the cell.
These pathways of MHC-restricted antigen presentation ensure that most of the body’s cells are screened for the possible presence of foreign antigens. The pathways also ensure that proteins from extracellular microbes preferentially generate peptides bound to class II MHC molecules for recognition by CD4+ helper T cells, which activate effector mechanisms that eliminate extracellular antigens. Conversely, proteins synthesized by intracellular (cytosolic) microbes generate peptides bound to class I MHC molecules for recognition by CD8+ CTLs, which function to eliminate cells harboring intracellular infections. The immunogenicity of foreign protein antigens depends on the ability of antigen-processing pathways to generate peptides from these proteins that bind to self MHC molecules.
These pathways of MHC-restricted antigen presentation ensure that most of the body’s cells are screened for the possible presence of foreign antigens.
The immunogenicity of foreign protein antigens depends on the ability of antigen-processing pathways to generate peptides from these proteins that bind to self MHC molecules.
