ANTIGEN PRESENTATION

Lymphocyte subsets Activate B cells and macrophages T HELPER CELLS Th Kill virus- infected cells CYTOTOXIC T LYMPHOCYTES CTL Produce antibodies PLASMA CELLS PC T B T CELLS B CELLS CLP Common lymphoid precursor

RECOGNITIONEFFECTOR CELL Macrophage activation Lymphocyte activation Inflammation Plasma cell Antibody production cytokines B-lymphocyte cytokines BCR + antigen Cytotoxic T- limfocyte (Tc) TCR + peptide + MHC-I Helper T- lymphocyte (Th) TCR + peptide + MHC-II Cell killing Effector cell retains specific receptor Effector cells secrete cytokines

Peptides of endogenous proteins (virus, tumor) bind to class I MHC molecules Tc Endogenous Ag RECOGNITION OF EXOGENOUS AND ENDOGENOUS ANTIGENES BY T-LYMPHOCYTES Exogenous Ag Th Peptides of exogenous proteins (toxin, bacteria, allergen) bind to class II MHC molecules

How can 6 invariant molecules have the capacity to bind to 1,000,000,000,000,000 different peptides? The number of different T cell antigen receptors is estimated to be 1,000,000,000,000,000 ( )

A flexible binding site? At the cell surface, such a binding site would be unable to with the T cell antigen receptor allow a high enough binding affinity to form a trimolecular complex with the T cell antigen receptor prevent exchange of the peptide with others in the extracellular milieu A binding site that is flexible enough to bind any peptide?

A 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. FloppyCompact Allows 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 Venus fly trap

MHC molecules Adopt a flexible “floppy” conformation until a peptide binds Fold around the peptide to increase stability of the complex The captured peptides contribute to the stabilization of the complex Use a small number of anchor residues to tether the peptide - this allows different sequences between anchors and different lengths of peptides WHERE PEPTIDE BINDING OCCURS?

INTRACELLULAR COMPARTMENTS ISOLATED BY MEMBRANSE 1) cytosol 2) vesicular system

THE ENDOGENOUS ANTIGEN PROCESSING PATHWAY Tc-cell Proteasome LMP2/LMP7 PROTEIN SELF ANTIGEN TAP1/2 gp96 calnexin α-chain α-chain+β2mMHC+peptide MHC-I + Ag peptide MHC-I + self peptide CLOSEDFLEXIBLE cytoplasm MHC-I, LMP2/7, TAP IFN  induced coordinated expression

CYTOSOL-DERIVED PEPTIDES ARE PRESENTED BY MHC-I FOR T-CELLS

Degradation of endogenous proteins in (immune) proteasomes TAP: Transporter associated with antigen processing

ER membrane Lumen of ER Cytosol Transporters associated with antigen processing (TAP1 & 2) Transporter has preference for longer than 8 amino acid peptides with hydrophobic C termini. TAP-1 TAP-2 Peptide TAP-1 TAP-2 Peptide TAP-1 TAP-2 Peptide TAP-1 TAP-2 Peptide TAP-1 TAP-2 Peptide TAP-1 TAP-2 Peptide TAP-1 TAP-2 Peptide TAP-1 TAP-2 Peptide TAP-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 Hydrophobic transmembrane domain Peptide antigens from proteasome

PEPTIDE-MHC INTERACTION IS SUPPORTED BY MULTIPLE PROTEINS IN THE ENDOPLASMIC RETICULUM

TRIMMING OF PEPTIDES FOR OPTIMAL SIZE BY BY ERAP

Th-cell INVARIANT CHAIN (Ii) 1.Chaperone – conformation 2.Inhibition of peptide binding 3.Transport/retention CLOSEDFLEXIBLE Ii+αβ CLIP DMA/B DMA/DMB 1. Support the peptide receptive conformation 2. Exchange of CLIP for exogenous peptides MHC-II + Ag peptide MHC-II + self peptide THE EXOGENOUS ANTIGEN PROCESSING PATHWAY

THE INVARIANT CHAIN PROTECTS THE MHC CLASS II BINDING SITE UNTIL REACHING THE APPROPRIATE COMPARTMENT INVARIANT CHAIN LÁNC (Ii) 1.Chaperon – Conformation 2.Inhibition of the peptide binding site 3.Transport and retention DMA/DMB 1. Stabilization of peptide accessable conformation 2. Exchange of CLIP to peptides derived from exogenous proteins

GENERATION OF MHC – II EPITOPESGENERATION OF MHC – I EPITOPES Viral protein B27 A2 B35 C42 HLA-A,B,C binding The Tc response is focused to few epitopes ENSURE RECOGNITION OF MANY PATHOGENIC PROTEIN The Th response is directed to overlapping epitopes ENSURE RECOGNITION OF ALL PROTEINS HLA-DR1/HLA-DR4 Overlapping peptides HLA-DQ2/HLA-DQ7

TARGETS OF EPSTEIN-BARR VIRUS-SPECIFIC Tc (CTL) RESPONSES A poliklonális CTL válasz elsősorban a litikus antigének és az A poliklonális CTL válasz elsősorban a litikus antigének és az EBNA3,4,6 nukleáris fehérjék ellen irányul EBNA3,4,6 nukleáris fehérjék ellen irányul Erősen fókuszált egy adott MHC - peptid kombinációra Erősen fókuszált egy adott MHC - peptid kombinációra Az endogén EBNA1 nem processzálódik és így nem ismerhető fel Az endogén EBNA1 nem processzálódik és így nem ismerhető fel EBNA3 EBNA5EBNA2 WWWWWWWWWWWW ZZ WWWWWWWWWW CC YY HHFFQQUUPPOOMMSSLLEERRKKBBDDTTXXVVIIAAGGNN hhee tt NN hh eettEBNA6 EBNA1LMP2 EBNA4 LMP1 - BZLF1BMLF1BMRF1BHRF1BARF0 + ± (?) ± LYTIC ANTIGENS LATENT ANTIGENS

MHC class IMHC class II Bound peptide sourceself or foreign proteins size8-10 amino acids13-21 amino acids heterogenitylimitedoverlapping set of peptides naturalcytoplasmic and nuclear proteins~70% MHC derived, membrane and extracellular proteins Generation of peptide sitecytoplasmvesicles, endo/lysosomes enzymeproteasome LMP-2, LMP-7 regulatory subunits vesicular acidic proteases cathepsins transportTAP - size and C-terminal dependent cytoplasm  ER no MHC transport noIi - target, retention ER  vesicular system special compartment MHC - peptide interaction siteERvesicles, CIIV chaperonscalnexin, toposinIi - CLIP, DMA/B MHC - peptide complexes In the cell surface stable complexes reflecting the endogenous environment of the cell few instable empty molecules stable complexes reflecting the exogenous/endogenous environment of the cell few re-circulating molecules complexed with CLIP

REGULATION OF CLASS I AND II MHC MOLECULES IFNγ IFNγR Type II immune IFNγ increases MHC expression Inflammatory cytokines and IFNγ induces MHC class II expression in certain tissue cells (endothelial, astrocyte, microglia) Co-ordinated upregulation of MHC-I, TAP, LMP and MHC-II, DM, Ii

B-cellT-cell Appearance of antigen Soluble, particles Any cell surface molecule Cells carrying self MHC-peptide complexes Nature of the antigén Natíve proteins, carbohydrate, lipids, metals, any structure Processed protein fragments = peptides Ligand Conformational determinant Ssequential determinant MHC-peptide complex Antigen recognizing receptor on the cell surface Variable BCR ligand (antigen) – spcific bivalent Variable TCR MHC + peptide pecific monovalent Soluble antigen recognizing receptor antibody- Collaboration of other cells -Antigen processing and presenting cells APC – interaction of two cells Antigen processing, presentation -Intracellular enzymatic degradation, peptide or MHC transportation Result of full activation Production of effector molecule antibody = soluble BCR Activation of new genes Activation molecules, production of lymphokines, TCR on the cell surface Possibililties of cell activation FULL plasma cell, antibody PARTIAL funcional anergy APOPTOSIS FULL Various lymphokines PARTIAL functional anergy certain lymphokines APOPTOSIS Co-receptors CD19, CD21,CD4, CD8, CD28/CTLA4, CD2, CD38