2REQUIREMENTS FOR ANTIGEN PRESENTATION Expression of MHC moleculesANTIGENa) Synthesis of viral antigens - intracellularb) Uptake of protein antigens – extracellular3. „Processing” of antigengeneration of peptides suitable for T-cell recognition4. Presentation of peptides in complex with MHC molecules on the cell surfaceT-cells with TCR are specialized for recognizing protein – derived fragments
4MOLECULES CONTAINING ONE OR MORE Ig DOMAIN(S) MEMBERS OF THE IMMUNOGLOBULIN SUPERGENE FAMILYMOLECULES CONTAINING ONE OR MORE Ig DOMAIN(S)V or C domain relatedFUNCTIONRECOGNITIONIg, TCR, MHC-I, MHC-IIADHESIONICAM-1, ICAM-2, VCAM-1, NCAMBINDINGCD4, CD8, CD28, B7, IL-1RI, PDGFR, FcRII, poly-IgR
5There are two types of MHC molecule, MHC class I and MHC class II. The two classes of MHC molecule have similar overall three-dimensional structures. Where they differ is in their constituent polypeptide chains. An MHC class II molecule is made of two similarly sized polypeptides that contain two extracellular domains and are anchored in the plasma membrane. In the MHC class I molecule, a larger polypeptide contains three extracellular domains and is anchored in the plasma membrane; a smaller polypeptide comprises the fourth extracellular domain and is not attached to the membrane.
6CELL SURFACE EXPRESSION OF MHC ON VARIOUS CELL TYPES Szövet MHC I MHC IIT cells /-B cellsMakrophagesDendritic cellsEpithelial cellsNeutrophylsHepatocytesKidneyBrainEritrocytesCell surface expression of MHC is influenced by activationMHC class I molecules are important in immune responses agains viruses and tumour cellsMHC class II plays a role in the activation of immunocytes and in the regulation of cell cell cooperation
8EGY VAGY TÖBB Ig DOMÉNT TARTALMAZÓ FEHÉRJÉK V vagy C doménhez hasonló Some members of the immunoglobulin supergene familyEGY VAGY TÖBB Ig DOMÉNT TARTALMAZÓ FEHÉRJÉKV vagy C doménhez hasonlóFUNKCTIONRECOGNITIONIg, TCR, MHC-I, MHC-IIADHESIONICAM-1, ICAM-2, VCAM-1, NCAMBINDINGCD4, CD8, CD28, B7, IL-1RI, PDGFR, FcRII, poly-IgR
9MHC class I and MHC class II molecules bind to different T-cell co-receptors. Left panel: the CD8 co-receptor of a CD8 T cells binds to an MHC class I molecule on an antigenpresenting cell (APC). Right panel: the CD4 co-receptor of a CD4 T cell binds to an MHC class II molecule on an APC. TCR, T-cell receptor. Although CD4 and CD8 perform an analogous function they have different polypeptide chains and three-dimensional structures.
10The structure of MHC proteins An MHC class I molecule (left panels) is composed of one membrane-bound heavy (or α) chain and noncovalently bonded β2-microglobulin. The heavy chain has three extracellular domains, of which the amino-terminal α1 and α2 domains resemble each other in structure and form the peptidebinding site. An MHC class II molecule (right panels) is composed of two membrane-bound chains, an α chain (which is a different protein from MHC class I α) and a β chain. These have two extracellular domains each, the aminoterminal two (α1 and β1) resembling each other in structure and forming the peptide-binding site. The β2 domain of MHC class II molecules should not be confused with the β2-microglobulin of MHC class I molecules. The ribbon diagrams in the lower panels trace the paths of the polypeptide backbone chains.
11THE PEPTIDE BINDING SITE OF MHC CLASS I MOLECULES
12THE PEPTIDE BINDING SITE OF MHC CLASS II MOLECULES
13Cleft geometry b2-M a-chain Peptide a-chain b-chain Peptide MHC class I accommodatepeptides of 8-10 amino acidsMHC class II accommodatepeptides of >13 amino acids
14The number of different T cell antigen receptors is estimated to be 1,000,000,000,000,000 (1015)How can 6 invariant molecules have the capacity tobind to 1,000,000,000,000,000 different peptides?
15Peptides can be eluted from MHC molecules Acid elute peptides
16Eluted peptides from MHC molecules have different sequences but contain motifs Peptides bound to a particular type of MHC class I molecule have conserved patterns of amino acidsA common sequence in a peptide antigen that binds to an MHC molecule is called a MOTIFRTYQLVNCPEIYSFHAmino acids common to many peptides tether the peptide to structural features of the MHC moleculeANCHOR RESIDUESAVTYKQLPSAYIKTethering amino acids need not be identical but must be relatedY & F are aromaticV, L & I are hydrophobicRGYVQLSIFNEKLAPGYNLSide chains of anchor residues bind into POCKETS in the MHC moleculeDifferent types of MHC molecule bind peptides with different patterns of conserved amino acids
18A flexible binding site? A binding site that is flexible at an early, intracellular stage of maturation Formed by folding the MHC molecules around the peptide.Venus fly trapFloppyCompactAllows a single type of MHC molecule to• bind many different peptides• bind peptides with high affinity• form stable complexes at the cell surface• Export only molecules that have captured a peptide to the cell surface
19MHC molecules bind peptides according to the following principals • Use a small number of anchor residues to tether the peptide- this allows different sequences between anchorsand different lengths of peptide• Adopt a flexible “floppy” conformation until a peptide binds• Fold around the peptide to increase stability of the complex
21THE ENDOGENOUS AND EXOGENOUS ROUTES OF ANTIGEN PRESENTATION Endogenous peptides are presented on MHC I (vírus proteins, tumor antigens)TcThExogenous peptides (toxins, bacterium, allergen) are presented by MHC IIExogenous AgEndogenous Ag
22The MHC I receptor binds the CD8 receptor, while MHC II binds CD4.The CD8 co-receptor binds to the α3 domain of the MHC class I heavy chain, ensuring that MHC class I molecules present peptides only to CD8 T cells (left panel). In a complementary fashion, the CD4 co-receptor binds to the β2 domain of MHC class II molecules, ensuring that peptides bound by MHC class II stimulate only CD4 T cells (right panel).
23Allelic polymorphism is concentrated in the peptide antigen binding siteClass IClass II(HLA-DR)1322m2121Polymorphism in the MHC affects peptide antigen bindingAllelic variants may differ by 20 amino acids
24Cytosol-derived peptides are presented by MHCI receptors
25Degradation of endogenous proteins in the (immun)proteosomesTAP: Transporter associatedWith antigen processingIn all cells, proteasomes degrade cellular proteins that are poorly folded, damaged, or unwanted. When a cell becomes infected, pathogen-derived proteins in the cytosol are also degraded by the proteasome. Peptides are transported from the cytosol into the lumen of the endoplasmic reticulum by the protein called transporter associated with antigen processing (TAP), which is embedded in the endoplasmic reticulum membrane.
26Multiple proteins help Ag presentation of MHCI MHC class I heavy chains assemble in the endoplasmic reticulum with the membrane-bound protein calnexin. When this complex binds β2-microglobulin (β2m) the partly folded MHC class I molecule is released from calnexin and then associates with the TAP, tapasin, calreticulin, ERp57, and protein disulfide isomerase (PDI) to form the peptide-loading complex. The MHC class I molecule is retained in the endoplasmic reticulum until it binds a peptide, which completes the folding of the molecule. The peptide:MHC class I molecule is then released from the other proteins and leaves the endoplasmic reticulum for transport to the cell surface.
27Trimming of antigenic peptides by ERAP The endoplasmic reticulum aminopeptidase (ERAP) binds to MHC class I molecules in which the amino terminus of an overlong peptide hangs out of the binding site. It removes the accessible amino acid residues to leave a peptide of 8-10 amino acids with an improved fit to the heterodimer of the class I heavy chain and β2-microglobulin. It is not known whether ERAP acts on the class I molecule alone, as illustrated here, or when it is part of the peptide-loading complex, or both.
28Presentation of extracellular (Exogen) peptides bemutatása (MHCII prezentáció)Processing of antigens for presentation by MHC class II or MHC class I molecules occurs in different cellular compartments.The left half of the figure shows the fate of peptides derived from extracellular antigens and pathogens. Extracellular material is taken up by endocytosis and phagocytosis into the vesicular system of the cell, in this case a macrophage. Proteases in these vesicles break down proteins to produce peptides that are bound by MHC class II molecules, which have been transported to the vesicles via the endoplasmic reticulum (ER) and the Golgi apparatus. The peptide:MHC class II complex is transported to the cell surface in outgoing vesicles. The right half of the figure shows the fate of peptides generated in the cytosol as a result of infection with viruses or intracytosolic bacteria. Proteins from such pathogens are broken down in the cytosol by the proteasome to peptides, which enter the ER. There the peptides are bound by MHC class I molecules. The peptide:MHC class I complex is transported to the cell surface via the Golgi apparatus.
29Invariant chain protects the binding site of MHCII until it reaches the appropriatecompartmentThe invariant chain prevents peptides from binding to an MHC class II molecule until it reaches the site of extracellular protein breakdown.In the endoplasmic reticulum (ER), MHC class II α and β chains are assembled with an invariant chain that fills the peptide-binding groove; this complex is transported to the acidified vesicles of the endocytic system. The invariant chain is broken down, leaving a small fragment called class II-associated invariant-chain peptide (CLIP) attached in the peptide-binding site. The vesicle membrane protein HLA-DM catalyzes the release of the CLIP fragment and its replacement by a peptide derived from endocytosed antigen that has been degraded within the acidic interior of the vesicles.INVARIÁNS LÁNC (Ii)Chaperon – konformációPeptidkötőhely gátlásaSzállító/visszatartó molekulaDMA/DMB1. A peptidet befogadó konformáció fenntartása2. A CLIP és az exogén fehérjékből származó peptidek lecserélése
31AZ MHC FUNCTIONSPresentation of peptides– self/altered self/foeign peptidesContinous expression of self-peptidesto monitor cellular healthDetermination of immunological selfSelf MHC + self peptide – individual MHC pluss és saját peptidAllogeneic response to fotreign MHC (transplantation)Self MHC– autolgous foreign MHC allogeneic activation. Az The ratio of alloreactive T-cells is very high: 1-10%A differentiation and selection of developing thymocytes (in the thymus)promotion of T-limphocyte survival in the pripheryweek” tonic” signals induced by MHC / TCR interactions provide survival signalsInhibitory ligands for NK cells, maintainance of host cell integrity.
32AZ MHC restriction TCR/ MHC + peptid complex recognized A single TCR recognize a single MHC-peptid komplexThe same peptide presented on a different MHC is not recognized.The same MHC molecule with a different peptide is not recognized by a given TCR (other TCRs may recognize)
33Experiment of Peter DOHERTY & Rolph ZINKERNAGEL 1976 Vírus B+ Y sejtekTVirus AT - CELLSTMICE YVírus A+ Y sejtekTMICE XVírus A+ X cellsTVirus A+ X cellsTTion)Cells infected with a virus are only killed if the infected cell and virus-specific T cells are from the same animal or strain. (The MHC needs to be recognized by the CTL cells MHC restriction ).MediaGraphicsInternational
34No T cells No rejection MiceY Thymectomy Mice X MiceX Tissue compatibility is encoded by the MHC genes and tissue rejection requires the presence of T cellsMiceYMiceXNo rejectionMice XThymectomyNo T cells
35MHC protein (HLA)- coding genes The MHC locusMHC protein (HLA)- coding genes
36The full sequence and the map of the human MHC locus HUMAN GENOME PROJECT3,838,986 bp224 gene6 kromoszómaMHC sequencing consortiumNature 401, 1999
37STRUCTURE OF THE MHC Non- classical MHC genes 6 kromoszóma rövid karja MHC15 kromoszóma 2mNon- classical MHC genesE, G, FKlasszikus MHC gének POLIMORPHICHLA – Human Leukocyte Antigen systemHLA –A, B, C I osztályALL NUCLEATED CELLLSHLA – DR, DP, DQ Class IION PROFESSIONAL APCClass III
38Of the human MHC class I isotypes, HLA-A, HLA-B, and HLA-C are highly polymorphic. They present peptide antigens to CD8 T cells and also interact with NK-cell receptors. HLA-E and HLA-G are oligomorphic and interact with NK-cell receptors. HLA-F is intracellular and of unknown function, and occurs as a single isotype. Of the human MHC class II isotypes, HLA-DP, HLA-DQ, and HLA-DR are polymorphic and present peptide antigens to CD4 T cells, whereas HLA-DM and HLA-DO occur in only a few isotypes, are intracellular, and regulate the loading of peptides onto HLA-DP, HLA-DQ, and HLA-DR.
39INHERITENCE OF CLASS I AND CLASS II MHC GENES HUMAN LEUKOCYTE ANTIGEN HLAKo-domináns öröklésmenetI osztályII osztályEVERY CELLα1β1α2β2PROFESSIONAL APC
41A polimorfizmus (allélek) száma ~6 x 1015 combinationsPOLIMORPHYSM OF MHC IN HUMAN POPULATIONS872CLASS I1652 allels50615.1828.6513.384.460.025.7218.888.449.921.884.4824.632.641.760.01CAUAFRASIFrequency (%)HLA-A1HLA- A2HLA- A3HLA- A28HLA- A36Allels274A polimorfizmus (allélek) számaABCCLASS II688 allels4661146615252In reality allels are not inherited randomy. Allels are linked, and there must have been strong selection favoring certain allelic variants. Non-random distribution.abDRDPDQ
42Classical MHC genes are inherited as haplotypes BCADPDQDROffspringDP-1,8DQ-3,6DR-5,4B-7,2C-9,8A-11,10DP-1,9DQ-3,7DR-5,5B-7,3C-9,1A-11,9DP-2,8DQ-4,6DR-6,4B-8,2C-10,8A-12,10DP-2,9DQ-4,7DR-6,5B-8,3C-10,10A-12,9BCADPDQDRXParentsBCADPDQDRDP-1,2DQ-3,4DR-5,6B-7,8C-9,10A-11,12DP-9,8DQ-7,6DR-5,4B-3,2C-1,8A-9,10BCADPDQDRBCADPDQDR
43Kidney epithelial cell B-cell, macrophage, dendritic cell MHC MOLECULES ARE EXPRESSED WITH BOUND PEPTIDES DERIVED FROM SELF OR NON-SELF PROTEINSKidney epithelial cellB-cell, macrophage, dendritic cellPresent intra- and extracellular environmentLiver cellClass II MHCOverlapping peptides of various sizes, which derive from membrane proteins70% derives from MHC moleculesPresent intracellular environmentClass I MHCPeptides of restricted size, which derive from cytosolic or nuclear proteins
44MHC Polimorphysm is maintained by the presence of pathogens In our example population there are four different MHC haplotypes, each represented by a different color. The frequencies of different genotypes are represented by the 20 circles in each panel, the frequencies of the four haplotypes being given underneath. First panel: the population experiences a period characterized by balancing selection arising from successive epidemic infections, after which only heterozygotes survive (second panel) and in which 30% of the population die, as indicated by circles containing an X. After recovery of the population during a period of relative calm and health (third panel) it becomes subject to directional selection by a new and particularly nasty infection. Only individuals with the blue MHC haplotype survive and 75% of the population dies (fourth panel). As a result of these selections the frequencies of the MHC haplotypes change considerably, but all four MHC haplotypes are retained within the population.Figure 5.35 MHC heterozygosity delays the progression to AIDS in people infected with HIV-1.When people who have been infected with HIV-1 start to make detectable antibodies to the virus they are said to have undergone seroconversion. The onset of overt symptoms of AIDS occurs years after seroconversion. The rate of progress to AIDS decreases with the extent of HLA heterozygosity, as shown here by a comparison of individuals who are heterozygous for all the highly polymorphic HLA class I and II loci (red) with those who are homozygous for one locus (yellow) or for two or three loci (blue).
45THE OUTCOME OF INFECTION IN A POPULATION WITH POLYMORPHIC MHC GENES Example: If MHC X was the only type of MHC moleculeMHC-GenvPathogen that evades MHC XMHCXXPopulation threatenedwith extinctionV – virus infectionPopulation is protected