Chapter 11

Antigen Presenting Cells and

Antigen Presenting

T cells do not recognise native antigens

YY Y Y YYY

BY

T

Y

T

Proliferation and antibody production

No proliferationNo cytokine release

Cross-linking of surface membrane Ig

Y

B

Y

B Y

B

Y

B

Y

B Y

B

Y

BYY

B

Cell surfacepeptides

of Ag

Antigens must be processed in orderto be recognised by T cells

YT

T cellresponse

No T cellresponse

No T cellresponse

No T cellresponse

No T cellresponse

Solublenative Ag

Cell surfacenative Ag

Soluble peptides

of Ag

Cell surface peptides of Ag presented by cells that

express MHC molecules

ANTIGENANTIGENPROCESSINGPROCESSING

APCAPC

Contents

Part Introduction--conceptsⅠ Part Characteristics of APCs Ⅱ Part Ag Processing and presentation Ⅲ

Part Introduction--conceptsⅠEndogenous Ags: antigens synthesized within cells, including

self and unself protein----class MHC molecules.ⅠExogenous Ags: antigens comes outside the cells, including

self and unself protein----class MHCⅡ molecules.

Antigen processing: the conversion of native proteins to peptides which can combine with MHC molecules.

Antigen presentation: the course of formation and display of

peptide-MHC complexes on the surface of APCs and the course of peptide-MHC complexes recognition by T cells.

Ag capturing----Endocytosis (internalization)

Phagocytosis, Pinocytosis, Receptor-mediated endocytosis

Production of endogenous Ags and exogenous Ags

YThe site of pathogen replication or mechanism of antigen uptake

determines the antigen processing pathway used

Y

Cytosolic compartmentEndogenous processing(Viral, tumor antigens )

Vesicular CompartmentContiguous with extracellular fluid

Exogenous processing(Streptococcal, tumor antigens)

INTRACELLULAR REPLICATION

EXTRACELLULAR ORENDOSOMAL REPLICATION

Cell surfacepeptides

of Ag

Antigens must be processed in orderto be recognised by T cells

YT

T cellresponse

No T cellresponse

No T cellresponse

No T cellresponse

No T cellresponse

Solublenative Ag

Cell surfacenative Ag

Soluble peptides

of Ag

Cell surface peptides of Ag presented by cells that

express MHC antigens

ANTIGEN PROCESSING

APCAPC

Antigen-Presenting Cells (APC)

APC (Accessory cells) : A group of cells play important roles in the immune response which can uptake, process antigens and present peptide-MHC complexes to T cells.

Professional APC: express classⅡMHC molecules Dendritic cell Macrophage B lymphocyte Facultative APC: endothelial cells, epithelial cells,

fibroblast, etc

APC

• Express class , MHC moleculesⅠ Ⅱ and co-stimulatory molecules

• Uptake, process endogenous/exogenous antigens and present peptide-MHC to T cells

• Including dendritic cells, macrophages and B cells

Part Characteristics of APCsⅡ

Dendritic cell (DC) Macrophage B lymphocyte

1. Dendritic cell (DC)

• History: DCs were first found by Steinman in 1973 ， named for their special spinelike projections. DCs were cultured successfully in vitro in 1993 by Inaba.

• Characteristic: The most efficient APC, can present antigens to naive T cells to elicit primary immune response.

Fig2-4 Mature DC suspended in media by colony （ ×400 ）

Fig2-5 scattered mature DC （ ×400 ）

Scanning electron micrograph

1. Dendritic cell (DC)

(1) Identification of DC: Typical morphology—spinelike projection MLR—stimulate naïve T cells activation Surface markers : CD1a, CD11c, CD83(human) high expression of classⅡMHC co-stimulatory molecules--CD80,CD86 others—CKs, CAMs, R(2) Source of DC: pluripotent hematopoietic stem cells myeloid DC myeloid progenitor lymphoid DC lymphoid progenitor

GM-CSF, IL-4

1. Dendritic cell (DC)

(3) Classification of DC : DC in lymphoid tissue:

Interdigitating DC (IDC) ,

Folicular DC (FDC) DC in non lymphoid tissue:

Langerhans cell (LC) DC in body fluid: Veiled cell, Blood DC

　　　 Interdigitating DC( IDC ）

Express high level of class , MHC molecules and B7Ⅰ Ⅱ ， lack of FcR and CR, can stimulate T cells.

FDC

B cell

　 Folicular DC （ FDC ）

Lie in follicle of LN, no expression of class ⅡMHC, high level of FcR and C3bR.

Langerhan’s cells(LC)—Birbeck particle

Lie in the epithelia of the skin, gastrointestinal and respiratory tracts, express FcR and C3bR. After uptaking antigens, migrating to draining LN and becoming IDC.

1. Dendritic cell (DC)

(4) Development and Maturation of DC

Pre-DC phase

Immature DC （ iDC ） phase

Migration phase

Mature DC （ mDC ） phase

Pre-DCBlood

Non-lymphoid tissueDifferentiation

ImmatureDC

DistributeWidely distributed in the body

Possess ability of Ag capture and process

Cytokines and Ag

DC mature and move into lymphoid tissue

Ability of Ag capture and processing decreases while its ability of Ag presenting increases

Difference between iDC and mDC

• Ability of uptaking and processing antigens decreases.

• Ability of antigen presentation increases.• Express high level of MHC, co-stimulatory

molecules(CD80,CD86), CAMs(ICAM-1).• Ability to stimulate naïve T cell activation

increases.

1. Dendritic cell (DC)

(5) Antigens capturing:

• Phagocytosis—cell, bacteria

• Pinocytosis—soluble antigen

• Receptor-mediated endocytosis

FcγRⅡ, C3bR, mannose receptor

(6) Function of DC :

• Capture, process, present antigens—APC

• Stimulate T or B lymphocytes—mature DC

• Induce immune tolerance—immature DC

1. Dendritic cell (DC)

2. Macrophage （ MФ）• Stem from monocytes in blood

• Have strong phagocytosis (big phagocyte)

• Can not stimulate naïve T cells

• Capture antigens by phagocytosis, pinocytosis, receptor-mediated endocytosis

Function : • Phagocytosis

• Presentation of antigens Unactivated macrophage

Activated macrophage:

Class MHC molecules Ⅱ and co-stimulatory molecules

2. Macrophage （ MФ）

3. B cells

Functions

• Mediate humoral immune response

• Immunological regulation

• Present antigens to T cell

Soluble Ag--pinocytosis

Specific receptor-mediated endocytosis

The three kinds of professional APC

Cell surfacepeptides

of Ag

Antigens must be processed in orderto be recognised by T cells

YT

T cellresponse

No T cellresponse

No T cellresponse

No T cellresponse

No T cellresponse

Solublenative Ag

Cell surfacenative Ag

Soluble peptides

of Ag

Cell surface peptides of Ag presented by cells that express MHC antigens

ANTIGEN PROCESSING

APCAPC

Part Ag Processing and PresentationⅢ

Class MHC pathway ------exogenous antigensⅡ

Class MHC pathway ------endogenous antigensⅠ

Cross – presentation of antigen

Section Class MHC pathwayⅠ Ⅱ 1. Capture of exogenous Ag

2. Processing of Ag

3. Synthesis and transportation of class Ⅱ MHC molecules

4. Peptide loading of classⅡ MHC molecules

5. Presenting to CD4+T cells

1. Capture of exogenous Ag

• Endocytosis:

Phagocytosis: particles or granules

Pinocytosis: soluble antigens

Receptor-mediated endocytosis:

• Form endosome

Y Y

Pinocytosis

Phagocytosis

Membrane Igreceptor mediateduptake

Y

Uptake of exogenous antigens

Complement receptormediated phagocytosis Y

Fc receptor mediated phagocytosis

2. Processing of Ag

endosome + lysosome

Ag antigen peptides(10-30aa) Cathepsin

Endosomes

Exogenous pathway

Cell surface

To lysosomes

UptakeProtein antigens

In endosome

Cathepsin B, D and L proteases are activated by the decrease in pH

3. Synthesis and transportation of class MHC Ⅱmolecules

Synthesis of class MHC molecules in ERⅡ

Ii chain --- class MHC moleculeⅡ (Ii3α3β3 ) ①Promote formation of class Ⅱ MHC dimer ②Preventing endogenous peptide from combining with

classⅡMHC molecules within ER

③Leading classⅡMHC molecules into endosome from ER

Endosome (MIIC)

*Ii chain: Ia-associated invariant chain

A peptide of the invariant chain blocks the MHC molecule binding site.This peptide is called the CLass associated Ⅱ Invariant chain Peptide (CLIP)

Invariant chain CLIP peptide

and b chains of MHC class II molecules

CLIP

Need to prevent newly synthesised, unfolded self proteins from binding to

immature MHC

Invariant chain stabilises MHC class II by non- covalently binding to the

immature MHC class II molecule and forming a nonomeric complex

In the endoplasmic reticulum

MHC class II maturation and invariant chain

Ii c

hain

CLIP

4. Peptide loading of class MHC moleculesⅡ

Ii - class MHC moleculesⅡ

protease Ii chain cleaving

CLIP - class MHC moleculesⅡ

HLA-DM CLIP releasing

Antigen peptide - class MHC complexesⅡ

Endosomes

Cell surface

Uptake

Class II associated invariant chain peptide (CLIP)

(inv)3 complexesdirected towardsendosomes byinvariant chain

Cathepsin L degrades Invariant chain

CLIP blocks groove in MHC molecule

MHC Class IIcontaining vesiclesfuse with antigen

containing vesicles

Removal of CLIP

?

How can the peptide stably bind to a floppy binding site?

Competition between large number of peptides

HLA-DM catalyses the removal of CLIP

MIIC compartment

HLA-DM

Replaces CLIP with a peptide antigen using a

catalytic mechanism (i.e. efficient at sub-

stoichiometric levels)

Discovered using mutant cell lines that failed to

present antigen

HLA-DO may also play a role in peptide exchange

Sequence in cytoplasmic tail retains HLA-DM in endosomes

HLA-DMHLA-DR

5.Presenting to CD4+T cells

Antigen peptide-class MHC molecuelsⅡ presented on cell membrane by exocytosis

MIIC compartment sorts peptide-MHC complexes for surface expression orlysosomal degradation

Surface expression of class II MHC -peptide complexes

Exported to the cell surface (t1/2 = 50hr)

Sent to lysosomes for degradation

CD4+T cells

Section class Ⅱ MHC pathwayⅠ

1. Processing of endogenous Ag

2. Transporting of antigen peptide into ER

3. Peptide loading of class MHC moleculesⅠ

4. Presenting to CD8+T cells

1. Processing of endogenous Ag

• Proteosome : 20S, 26S

• Low molecular weight polypeptide (LMP) :

LMP2, LMP7 ， LMP10

• Ag antigen peptides (6-30aa)

Degradation in the proteasome

The components of the proteasome include LMP2, LMP7, MECL-1 （ LMP10 ）

*MECL-1 ： Multicatalytic endopeptidase complex subunit

Cytoplasmic cellular proteins, including non-self proteinsare degraded continuously by a multicatalytic protease

Binding ubiquitin

2. Transporting of antigen peptide into ER

TAP(transporter associated with antigen processing):

Consisting of TAP1 and TAP2 ATP dependent transporter

Selective transporting (8-15aa)

ENDOPLASMIC RETICULUM

CYTOSOL

Peptide antigens produced in the cytoplasm are physically separated from newly formed class I MHC

Newly synthesisedclass I MHC molecules

Peptides needaccess to the ER in

order to be loaded onto class I MHC molecules

ER membrane

Lumen of ER

Cytosol

Transporters associated withantigen processing (TAP1 & 2)

Transporter has preference for >8 amino acid peptideswith hydrophobic C termini.

TAP-1 TAP-2

Peptide

TAP-1 TAP-2

PeptideTAP-1 TAP-2

Peptide

TAP-1 TAP-2

PeptideTAP-1 TAP-2

Peptide

TAP-1 TAP-2

PeptideTAP-1 TAP-2

Peptide

TAP-1 TAP-2

PeptideTAP-1 TAP-2

Peptide

TAP-1 TAP-2

Peptide

ER membrane

Lumen of ER

Cytosol

TAP-1 TAP-2

Peptide

ATP-binding cassette(ABC) domain

Hydrophobictransmembranedomain

Peptide antigensfrom proteasome

3. Peptide loading of class MHC moleculesⅠ ER: antigen peptide—class MHC complexesⅠ

Endoplasmic reticulum

Calnexin bindsto nascent

class I chainuntil 2-M binds

TAP-1 TAP-2

Peptide

TAP-1 TAP-2

PeptideTAP-1 TAP-2

Peptide

TAP-1 TAP-2

PeptideTAP-1 TAP-2

Peptide

TAP-1 TAP-2

PeptideTAP-1 TAP-2

Peptide

TAP-1 TAP-2

PeptideTAP-1 TAP-2

Peptide

TAP-1 TAP-2

PeptideTAP-1 TAP-2

Peptide

B2-M binds and stabilises

floppy MHC

Tapasin, calreticulin, TAP 1 & 2 form a complex with

the floppy MHC

Cytoplasmic peptides are loaded onto the

MHC molecule and the structure becomes

compact

Maturation and loading of class I MHC

4. Presenting to CD8+T cells Antigen peptide-class Ⅰ MHC molecuels presented on cell membrane by exocytosis

Fate of class I MHC

Sent to lysosomes for degradation

Exported to the cell surface

Ag(cytosolic protein)

Proteasome proteolytic degradation

Ag peptide

TAP complex transporting into ER

antigen peptide-class Ⅰ MHC complexes

Golgi complex exocyotsis

Presenting to CD8+T cells

CD8+T cells

CD8+T cells

CD4+T cells

Section Cross-presentation of antigensⅢ

Cross-priming:

• Class MHC molecules also present exogenous Ⅰantigens to CD8+T cells

• Class MHC molecules also present endogenous Ⅱantigens to CD4+T cells

CD4+T cell(Th) CD8+T cell(Tc)

T cellReceptor

Peptide

MHCClass II

T cellReceptor

Peptide

MHCClass I

Antigen PresentingCell

Target cell

CD4 CD8