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complex

Cells that express

that MHC

Responsive T cells

Source of protein

antigens

Enzymes responsible

for peptide generation

Site of peptide

loading of MHC

Molecules involved in

transport of peptides

and loading of MHC

molecules

Polymorphic a chain of MHC,

ß2-microglobulin, peptide

All nucleated cells

CD8+

T cells

Cytosolic proteins (mostly

synthesized in the cell; may

enter cytosol from phagosomes)

Protease components of

cytosolic proteasome

Class I MHC pathway

Polymorphic a and ß chains

of MHC, peptide

Dendritic cells, mononuclear

phagocytes, B lymphocytes;

endothelial cells, thymic

epithelium

CD4+

T cells

Endosomal/lysosomal

proteins (mostly internalized

from extracellular environment)

Endosomal and lysosomal

proteases (e.g., cathepsins)

Endoplasmic reticulum Late endosomes

and lysosomes

TAP Invariant chain, DM

a ß2-microglobulin a ß

Peptide Peptide

Fig. 3.13 Features of the pathways of antigen processing. Some of the comparative features of the two

major antigen processing pathways. MHC, Major histocompatibility complex; TAP, transporter associated

with antigen processing.

66 CHAPTER 3 Antigen Capture and Presentation to Lymphocytes

becomes stable and is delivered to the cell surface.

If the MHC molecule does not find a peptide it can

bind, the empty molecule is unstable and is eventually degraded in the ER. One protein antigen may give

rise to many peptides, only a few of which (perhaps

only one or two from each antigen) can bind to the

MHC molecules present in the individual and have the

potential to stimulate immune responses in that individual. The class I MHC-peptide complexes are recognized by CD8+ T cells.

The evolutionary struggle between microbes and

their hosts is well illustrated by the numerous strategies that viruses have developed to block the class I

MHC pathway of antigen presentation. These strategies

include removing newly synthesized MHC molecules

from the ER, inhibiting the transcription of MHC genes,

and blocking peptide transport by TAP. By inhibiting

the class I MHC pathway, viruses reduce presentation of

their own antigens to CD8+ T cells and are thus able to

evade the adaptive immune system. These mechanisms

of immune evasion are discussed in Chapter 6.

Processing of Internalized Antigens for

Display by Class II MHC Molecules

The main steps in the presentation of peptides by class II

MHC molecules include internalization of the antigen,

proteolysis in endocytic vesicles, association of peptides

with class II molecules, and transport of peptide-MHC

complexes to the cell surface (Fig. 3.15).

Internalization and Proteolysis of Antigens

Antigens destined for the class II MHC pathway are

usually internalized from the extracellular environment. Dendritic cells and macrophages may ingest

Protein antigen

of ingested microbe

transported to cytosol ER

Exocytic

vesicle

Proteasome

Virus in

cytoplasm Synthesized

viral protein

Ubiquitinated

protein

Ub

Peptides

ß2m

TAP

Class I

MHC

a chain

Chaperone

Golgi

Phagosome

CD8

Surface

expression of

peptide-class I

complexes

Assembly of

peptide-class I

complexes

in ER

Transport

of peptides

from cytosol

to ER

Proteolytic

degradation

of proteins

Production

of proteins

in the

cytosol

Tapasin

CD8+

cytotoxic T

lymphocyte

Fig. 3.14 Class I MHC pathway of processing of cytosolic antigens. Proteins enter the cytoplasm of cells

either from endogenous synthesis by microbes, such as viruses, that reside in the cytosol (or nucleus, not

shown) of infected cells or from microbes that are ingested but whose antigens are transported into the cytosol (the process of cross-presentation, described later). Cytoplasmic proteins are unfolded, ubiquitinated, and

degraded in proteasomes. The peptides that are produced are transported by the transporter associated with

antigen processing (TAP) into the endoplasmic reticulum (ER), where the peptides may be further trimmed.

Newly synthesized class I MHC molecules are initially stabilized by chaperones and attached to TAP by a

linker protein called tapasin, so the MHC molecules are strategically located to receive peptides that are

transported into the ER by TAP. The peptide–class I MHC complexes are transported to the cell surface and

are recognized by CD8+ T cells. Ub, Ubiquitin; ß2m, ß2-microglobulin.

CHAPTER 3 Antigen Capture and Presentation to Lymphocytes 67

extracellular microbes or microbial proteins by several

mechanisms, including phagocytosis and receptormediated endocytosis. Microbes may bind to surface

receptors specific for microbial products or to receptors that recognize antibodies or products of complement activation (opsonins) attached to the microbes.

B lymphocytes efficiently internalize proteins that

specifically bind to the cells’ antigen receptors (see

Chapter 7). Certain APCs, especially dendritic cells,

may also pinocytose proteins without any specific

recognition event. After internalization into APCs by

any of these pathways, the microbial proteins enter

acidic intracellular vesicles, called endosomes or phagosomes, which fuse with lysosomes. In these vesicles,

the proteins are broken down by proteolytic enzymes,

generating many peptides of varying lengths and

sequences.

Binding of Peptides to Class II MHC Molecules

Peptides bind to newly synthesized class II MHC

molecules in specialized vesicles. Class II MHC–

expressing APCs constantly synthesize these MHC

molecules in the ER. Each newly synthesized class II

molecule carries with it an attached protein called the

invariant chain (Ii

), which contains a sequence called

the class II invariant chain peptide (CLIP) that binds to

the peptide-binding cleft of the class II molecule. Thus,

the cleft of the newly synthesized class II molecule is

occupied and prevented from accepting peptides in the

ER that are destined to bind to class I MHC molecules

(discussed earlier). This class II molecule with its associated Ii

 migrates from the ER through the Golgi stacks

and then, instead of traveling directly to the plasma

membrane, is targeted by the cytosolic tail of the invariant chain to acidic vesicles (endosomes and lysosomes).

Endocytic

vesicle

Endosome

Golgi

ER

Lysosome

HLA-DM

CLIP

CLIP

Class

II MHC

Exocytic

vesicle

CD4

CD4+

helper

T cell

Protein antigen

Uptake of

extracellular

proteins into

vesicular

compartments

of APC

Processing

of internalized

proteins in

endosomal/

lysosomal

vesicles

Biosynthesis and

transport of

class II MHC

molecules

to endosomes

Association

of processed

peptides with

class II MHC

molecules

in vesicles

Expression

of peptideMHC

complexes

on cell

surface

a

ß

Ii

Chaperone

Fig. 3.15 Class II major histocompatibility complex (MHC) pathway of processing of internalized vesicular antigens. Protein antigens are ingested by antigen-presenting cells (APCs) into vesicles, where they are

degraded into peptides. Class II MHC molecules enter the same vesicles, where the class II invariant chain peptide (CLIP) that occupies the cleft of newly synthesized class II molecules is removed. These class II molecules

are then able to bind peptides derived from the endocytosed protein. The DM molecule facilitates the removal

of CLIP and subsequent binding of the antigenic peptide. The peptide–class II MHC complexes are transported

to the cell surface and are recognized by CD4+ T cells. ER, Endoplasmic reticulum; Ii

, invariant chain.

68 CHAPTER 3 Antigen Capture and Presentation to Lymphocytes

In this compartment, the invariant chain is degraded,

leaving only CLIP in the peptide-binding cleft. Ingested

proteins are digested into peptides in the same compartment. The vesicles also contain a class II MHC–like

protein called DM, whose function is to exchange CLIP

in the class II MHC molecule with other peptides that

may be available in this compartment that can bind to

the MHC molecule with higher affinity.

Transport of Peptide-MHC Complexes to the Cell

Surface

Peptide loading stabilizes class II MHC molecules,

which are exported to the cell surface. If a class II molecule binds a peptide with the right fit, the complex is stabilized and transported to the cell surface, where it can be

recognized by a CD4+ T cell. Class II molecules that do

not find peptides they can bind are eventually degraded

by lysosomal proteases. As for the class I pathway, only

a few of the peptides produced from any protein antigen

can bind to MHC molecules present in the individual and

stimulate immune responses in each individual.

Cross-Presentation of Internalized Antigens

to CD8+ T Cells

Some dendritic cells can present ingested antigens on

class I MHC molecules to CD8+ T lymphocytes. This

pathway of antigen presentation is contrary to the general rule for APCs that most internalized proteins are displayed by class II MHC molecules to CD4+ T cells. The

initial response of naive CD8+ T cells, similar to CD4+

cells, requires that the antigens be presented by mature

dendritic cells in lymph nodes through which the naïve

T cells circulate. However, some viruses may infect only

particular cell types and not dendritic cells, and these

infected cells may not travel to lymph nodes or produce

all the signals needed to initiate T cell activation. How,

then, are naive CD8+ T lymphocytes in lymph nodes

able to respond to the intracellular antigens of infected

cells? Similarly, tumors arise from many different types

of cells, so how can diverse tumor antigens be presented

to naive CD8+ T cells in lymph nodes by dendritic cells?

A subset of classical dendritic cells has the ability to

ingest infected host cells, dead tumor cells, microbes,

and microbial and tumor antigens and transport the

ingested antigens into the cytosol, where they are processed by the proteasome. The antigenic peptides that

are generated then enter the ER and bind to class I molecules, which display the antigens for recognition by

CD8+ T lymphocytes (Fig. 3.16). This process is called

cross-presentation (or cross-priming), to indicate that

one type of cell, dendritic cells, can present the antigens

of other infected or dying cells or cell fragments and

Fragments

of cells and

antigens

picked up

by host APCs

Virally

infected

cell

Dendritic cell

Viral

antigen

Class II

MHC

Class I

MHC

Class I

MHC

Viral

antigen

enters

cytosol

Costimulator

Virus-specific

CD8+ T cell

Antigen

capture

Crosspresentation

T cell

response

Fig. 3.16 Class I MHC-restricted cross-presentation of microbial antigens from infected cells by dendritic cells. Fragments of cells infected with intracellular microbes (e.g., viruses) or antigens produced in

these cells are ingested by dendritic cells, and the antigens of the infectious microbes are broken down and

presented in association with class I MHC molecules of the antigen-presenting cells (APCs). T cells recognize the microbial antigens expressed on the APCs, and the T cells are activated. By convention, the term

cross-presentation (or cross-priming) is applied to CD8+ T cells (cytotoxic T lymphocytes) recognizing class I

MHC–associated antigens (as shown); the same cross-presenting APC may display class II MHC–associated

antigens from the microbe for recognition by CD4+ helper T cells.

CHAPTER 3 Antigen Capture and Presentation to Lymphocytes 69

prime (or activate) naive CD8+ T lymphocytes specific

for these antigens. Once the CD8+ T cells have differentiated into CTLs, they kill infected host cells or tumor

cells without the need for dendritic cells or signals other

than recognition of antigen (see Chapter 6). The same

pathway of cross-presentation is involved in initiating

CD8+ T cell responses to some antigens in organ transplants (see Chapter 10).

Physiologic Significance of MHC-Associated

Antigen Presentation

Many fundamental features of T cell–mediated immunity are closely linked to the peptide display function of

MHC molecules:

• The restriction of T cell recognition to MHC-associated

peptides ensures that T cells see and respond only to

cell-associated antigens. This is because MHC molecules are cell membrane proteins and because peptide

loading and subsequent expression of MHC molecules

depend on intracellular biosynthetic and assembly

steps. In other words, MHC molecules can be loaded

with peptides only inside cells, where intracellular and

ingested antigens are present. Therefore, T lymphocytes

can recognize the antigens of intracellular microbes,

which require T cell–mediated effector mechanisms, as

well as antigens ingested from the extracellular environment, such as those against which antibody responses

are generated.

• By segregating the class I and class II pathways of antigen processing, the immune system is able to respond

to extracellular and intracellular microbes in different ways that are specialized to defend against these

microbes (Fig. 3.17). Cytosolic antigens are processed

and displayed by class I MHC molecules, which are

expressed on all nucleated cells—as expected, because

all nucleated cells can be infected with one or more

species of viruses. Class I–associated peptides are recognized by CD8+ T lymphocytes, which differentiate

into CTLs. The CTLs kill the infected cells and eradicate the infection, this being the most effective mechanism for eliminating cytoplasmic microbes. CTLs

also kill tumor cells which produce cytosolic proteins from mutated genes. Many bacteria, fungi, and

even extracellular viruses are typically captured and

ingested by macrophages and their antigens are presented by class II molecules. Because of the specificity

of CD4 for class II, class II–associated peptides are

recognized by CD4+ T lymphocytes, which function

as helper cells. These T cells help the macrophages

to destroy ingested microbes, thereby activating an

effector mechanism that can eliminate microbes that

are internalized from the extracellular environment.

B lymphocytes ingest protein antigens of microbes

and also present processed peptides for recognition

by CD4+ helper T cells. These helper cells stimulate

the production of antibodies, which serve to eliminate extracellular microbes. Neither phagocytes nor

antibodies are effective against intracellular viruses

and other pathogens that can survive and replicate

in the cytoplasm of host cells; cells harboring these

cytosolic microbes are eliminated by CD8+ CTLs.

Thus, the nature of the protective immune

response to different microbes is optimized by linking several features of antigen presentation and T cell

recognition: the pathways of processing of vesicular

and cytosolic antigens, the cellular expression of class

I and class II MHC molecules, the specificity of CD8

and CD4 coreceptors for class I and class II molecules, and the functions of CD8+ cells as CTLs and

of CD4+ cells as helper cells. The function of linking

the type of microbe to one of the the two antigenprocessing pathways is important because the antigen

receptors of T cells cannot distinguish between intracellular and extracellular microbes. In fact, as previously mentioned, the same virus can be extracellular

early after infection and becomes intracellular once

the infection is established. During its extracellular

life, the virus is fought by antibodies and phagocytes,

whose production or functions are stimulated by

helper T cells, but once the virus has found a haven

in the cytoplasm of cells, it can be eradicated only by

CTL-mediated killing of the infected cells. The segregation of class I and class II antigen presentation

pathways ensures the correct, specialized immune

response against microbes in different locations.

• The structural constraints on peptide binding to

different MHC molecules, including length and

anchor residues, account for the immunodominance of some peptides derived from complex

protein antigens and for the inability of some

individuals to respond to certain protein antigens. When any protein is proteolytically degraded

in APCs, many peptides may be generated, but only

those peptides able to bind to the MHC molecules

in that individual can be presented for recognition

by T cells. These MHC-binding peptides are the

70 CHAPTER 3 Antigen Capture and Presentation to Lymphocytes

immunodominant peptides of the antigen. Even

microbes with complex protein antigens express

a limited number of immunodominant peptides.

Many attempts have been made to identify these

peptides in order to develop vaccines, but it is difficult to select a small number of peptides from

any microbe that would be immunogenic in a large

number of people, because of the enormous polymorphism of MHC molecules in the population. The

polymorphism of the MHC also means that some

individuals may not express MHC molecules capable of binding any peptide derived from a particular

antigen. These individuals would be nonresponders

to that antigen. One of the earliest observations that

established the physiologic importance of the MHC

was the discovery that some inbred animals did not

respond to simple protein antigens and responsiveness (or lack of) mapped to genes called immune

response (Ir) genes, later shown to be class II MHC

genes.

+

Antigen uptake

or synthesis

Antigen

presentation

T cell–dependent

effector functions

Class II MHC pathway:

antigen presentation to helper T cells

Macrophage

Extracellular

microbe

CD4+ helper

T lymphocyte

Microbe in

endosome

Macrophage activation:

killing of

phagocytosed microbe

B cell antibody

secretion: antibody

binding to antigen

Class I MHC pathway:

antigen presentation to cytotoxic T lymphocytes

Cytosolic

antigen CD8+ cytotoxic

T lymphocyte

Killing of

antigen-expressing

target cell