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Antigen processing and presentation

Monika Raulf

Lecture 25.04.2018

Monika Raulf - Antigen processing and presentation 2

AP is the display of peptide antigens (created via antigen processing) on the cell surface together with either MHC class I or class II molecules, which permit T cells to recognize antigens on the cell surface of an antigen-presenting cell (APC).

What is Antigen presentation ?


Which tools are necessary for antigen presentation?

What is antigen processing?


- APC - T cells


DC Macrophages B-cell Antigen presenting cells


Properties of different antigen presenting cells

lymphatic tissue, blood in the periphery

lymphatic tissue, conjunctive tissue, body cavities

lymphatic tissue, conjunctive tissue, epithelia

Localisation

soluble antigens, toxins, viruses

particular antigens, intra- and extra cellular

pathogens

peptides, viral antigens, allergens

Presented antigens

inducible; from − to +++

inducible; from − to +++

constitutive via mellow, not phagocytic, lymphatic

dendritic cells++++

Emission of co-stimulating signals

constitutive increase by activation;

from +++ to ++++

via bacteria and cytokines inducible;

from − to +++

on dendritic tissue cells low; on dendritic cells in lymphatic tissue significantly expressed

MHC-expression

antigen specific receptor (Ig) ++++

phagocytosis +++

+++ macro pino cytosis and phagocytosis by dendritic tissue cells; virus infection

Antigen exposure


from: „Immunologie“, Janeway et al.

Pathogens and their products are either in the cytoplasm or in the vesicles of the cell

Intra vesicular pathogens

Pathogens in cytosol

Extra cellular pathogens and toxins

Cytoplasm acidified vesicle acidified vesicle

MHC-class I MHC-class II MHC-class II

CD8 T-cells CD4 T-cells CD4 T-cells

Cell death

Activation for killing intra vesicular bacteria and

parasites

Activation of B-cells, Ig to secrete and

extra cellular bacteria or to eliminate toxins


Why is antigen presentation necessary?

Activation of T-cells* Viruses and some bacteria Parasites

Proliferate intracellular and are not accessible for antibodies

*T cells recognize antigens only in the context of self MHC molecules on the surface of accessory cells


Key player of the adaptive immunity

T-cells (origin is the thymus)

T-cells are essential for the control of intracellular viruses and for the activation of B-

cell-reactions against most antigens.


Cell-mediated reactions base on direct interactions between T-lymphocytes and cells, which support the T-cells to recognize an antigen.

T-helper cells [CD4⊕]

Cytotoxic cells [CD8⊕]

In both cases T-cells identify their target cells due to the peptide fragments, which originate from foreign proteins.


Human leukocyte antigen complex (HLA)

or

Major histocompatibility complex (MHC)

Are membrane surface glycoproteins used by T cells to recognize antigens

(structures necessary for the antigen-specific T cell activation)


MHC-Genes Important for die compatibility of tissue transplants

⇒ As several, closely connected, very polymorphic genes specify the histocompatibility, the concept „Histocompatibility complex“ is used


T-cells just recognize peptide fragments, which arise from an foreign antigen, if they are bound to MHC-molecules.

There are 2 classes of MHC-molecules:

MHC I-molecules

MHC II-molecules

⇒ Important distinguishing feature: Origin of the peptide, which is bound to the MHC-molecules and used it for the transport to the cell surface and the presentation


T-cell-peptide

MHC-molecule

T-cell

APC Proteases Antigen

1. Step: Antigen exposure/-processing


APC T-lymphocyte

TCR

CD28/ CTLA-4

2. signal

CD 80/86

1. signal

MHC+ peptide

2. Step: Stimulation of the T-cells by an antigen presenting cell Antigen


MHC = Major Histocompatibility Complex ⇒ Primary function: Binding and presentation of antigenic peptides on the cell surface for binding on antigen specific TCR on T cells.

MHC class I and class II have different structural features regarding the activation of variable T-lymphocytes

MHC I presents peptides for → cytotoxic Tc-lymphocytes

binds on

CD8 express

MHC II presents peptides for → helper TH-lymphocytes

binds on

CD4 express


Different antigen degeneration and processing ways, which lead to

⇒ „endogenous“ peptides associated with MHC I

⇒ „exogenous“ peptides associated with MHC II


MHC I: - endogenous synthesised antigens are proteolytic fragmented - small peptide fragments are transported to ER and bind with the arising (labile) MHC I-molecule - MHC I-peptide complex moves through the Golgi apparatus and reaches the cell surface

MHC II: - exogenous peptides, which are internalised from APC, are proteolytic fragmented - Peptide fragments are compartmentalised in the endosome and - associate to the MHC II-molecule; complex reaches the cell surface

⇒ Presentation → CD8-cells

⇒ Presentation → CD4-cells


The size of the antigen peptides, which reside in the „binding bag“ of MHC class I or class II, is different:

„MHC I-peptide“ → 8 – 10 AS

Different structures, as varying α- and β-polypeptide subunits exist, which maturate to αβ-heterodimer:

→ MHC I: α-UE (is coded of polymorphic MHC-genes) β-UE (not MHC-gene coded)

β2 microglobulin (β2m), very conserved → MHC II: α-UE

β-UE MHC-gene coded

αβ-dimer → tends to associate with (αβ)2

„MHC II-peptide“ → 13 – 18 AS


MHC binds peptides

Class I Class II


Differentiated cell specific expression of class I- and class II-molecules correlates with the specialisation of the immune cells → MHC I-expression is wide-spread on almost every somatic cell

cytotoxic T-cells have a protective function

→ MHC II-expression is exclusively on APC

local activation of helper T-cells

Important feature: Multi-peptide-binding


MHC class I- and class II- molecules

Structure of the MHC class I-molecule

Structure of the MHC class II-molecule

extra cellular domain*2

Constituted of α-chain + β2-microglobulin*1

*1 12 kD; not polymorph *2 belongs to the immunoglobulin superfamily


Genetic organisation of the main (major) histocompatibility complex in humans

HLA-locus (human leucocyte antigen) In humans on chromosome 6

MHC-molecules are polymorphic transmembrane glycoproteins

e.g. TAP 1, 2

β2-microglobulin (is part of the MHC-class I-molecules, but not located on chromosome 6)


Antigen processing and antigen presentation

MHC-molecules are instable without bound peptides; For MHC I-presentation important proteins/structures: • TAP1 + TAP2 (Transporters associated with antigen processing 1/2); TAP1:TAP2-complex is an ATP-dependent peptide transporter • Proteasome (multi catalytic protease complex) (Subunits of the proteasome (LMP2 + LMP7) are coded near the TAP1- and TAP2-genes in MHC; also their expression is inducible by IFN (IFN is virus-inducible)) • Proteins with chaperon-like functions (Calreticulin) („cellular quality management“) MHC I-class I-molecules leave the ER only, if they have bound peptides


Binding of the peptide (break up of proteins in the proteasome and transport via TAP into ER) to the complex induces the complete folding and the peptide-MHC complex leaves the ER and will be transported via Golgi-

apparatus to the cell surface

in ER: MHC I-α-chains store themselves together with calnexin (membrane-bound)

Binding of the MHC I-α/calnexin-complex to β2-microglobin (β2m)

New MHC I-αβ2m-dimer breaks away from calnexin

⇓

Complex in correlation with tapasin (is TAP-associated)

Chaperon molecules calreticulin + Erp57 contribute to the stability of the complex

MHC I-molecules exist in a cell abundantly, therefore in case of a viral infection the cell is able to react very fast

⇓

⇓

⇓

⇓


Extra-cellular proteins or pathogens, which reproduce intra-cellular (parasites, mycobacteria)

⇓ Affiliation in intra-cellular vesicle (endosomes)

⇓ Decline of the pH-value in the endosomes = Acidification effects the

activation of the proteases (acid proteases*) ⇓

Fusion of the endosomes during the migration into the cell interior with the lysosomes

⇓ Division of the protein antigens + binding on MHC II-molecules of the ER

are conducted by the invariant chain(Ii) to the acidified vesicles

Peptides, that are presented of MHC II-molecules, arise in acidified endocytotic vesicles

*e.g. Cysteine proteases as Cathepsin B, D and S as well as L


Antigen presentation via MHC II-molecules Important components: Invariant chain (Ii)

a) binds to the MHC II αβ-heterodimer; verifies the binding line for peptides

⇓ prevents the configuration by peptides in ER; in ER are the single components associated to calnexin ⇓

complete complex breaks away from calnexin and will be transported from the ER ⇒ Peptides of ER cannot be bound ⇓

b) provides for the transport of the complex to an endosomal compartment (probably MIIC = MHC II-compartment)

⇓ here Ii will be cut by proteases ⇓

it remains a short Ii-piece = CLIP


CLIP = class II-associated invariant chain peptide

Still eliminates the loading with the peptide

HLA-DM - binding and stabilisation of empty MHC II-molecules

- catalyses formation of the CLIP-fragment

- analogy to TAP-molecules at MHC I-molecules


The MHC-II way of presentation

Invariant chain with CLIP-fragment

Antigen

MHC-II

EE IE

Endosome

ER Antigen presenting cell

Golgi MIIC

Receptor mediated

Phagocytosis

proteolytic enzymes

EE = early endosomes IE = intermediate endosomes

ER = endoplasmatic reticulum MIIC = MHC-loaded compartment


Different features of the MHC-molecules MHC class I MHC class II

Expression upon all nucleated cells of the body

upon professional antigen presenting cells

Activation of CD8+ T-cells of CD4+ T-cells

Genloci HLA-A, HLA-B, HLA-C HLA-DR, HLA-DP, HLA-DQ

Structure Transmembrane constant α-chain associated with

β2-microglobulin

Transmembrane constant heterodimer of the α- and β-

chain

Peptides max. 8-10 amino acids long

at least 13 amino acids long

Loading in the endoplasmic reticulum in the intra-cellular vesicle


Peptides bound to MHC I-class stimulate CD8-cells

⇒ Strategies of viruses to prevent the mechanism of the antigen presentation: • Herpes simplex-virus: prevents the transport of viral peptide in the ER by the production of a protein that binds on the TAP-transporter and blockades it

• Adenovirus: synthesizes a protein, that can retain MHC I-molecules in the ER

• Cytomegalic virus precipitates the retrograde transport (HCMV): of MHC I-molecules back in the cytosol, where they are degraded


The role of T-effector cells in the cellular and humoral immune response against different disease agents

Cell-mediated immunity Humoral immunity

from: „Immunologie“, Janeway et al.

Typical pathogens

Vaccinia virus, Influenza virus, Rabies virus,

Listeria

Mycobacterium tuberculosis

Mycobacterium leprae Leishmania donovani, Pneumocystis carinii

Clostridium tetani Staphylococcus aureus

Streptococcus pneumoniae Poliovirus

Pneumocystis carinii

Localisation Cytosol Vesicle of macrophages

Extra cellular fluid

T-effector cell Cytotoxic

CD8-T-cell TH1-cell TH2/TH1-cell

Antigen recognition

Mode of action of the effector

Peptide: MHC-class-I on

infected cell

Peptide: MHC-class-II on

infected macrophages

Peptide: MHC-class-II on antigen

specific B-cell

Killing of the infected cell

Activation of infected macrophages

Activation of specific B-cells to synthesise

antibodies


Antigens, which are recognized by T-cells, have two different „Interaction areas“

Class II-MHC

Agretope

Epitope TCR

→ Agretope interacts with the MHC-molecule

→ Epitope interacts with the TCR

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