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Antigen recognition by T and B cellsRole of Antigen-Presenting Cells (APC)- T and B cells exhibit fundamental differences in- Helper T cells: recognize antigen after processing andpresentation by MHC-II on APC (dendritic cells,macrophages, B cells).antigen recognition- B cells recognize antigen free in solution (nativeantigen).- T cells recognize antigen after it has beenphagocytosed, degraded and small pieces of theantigen have been bound by MHC molecules.Antigen presenting cells- Cytotoxic T cells: recognize antigen when it is presented onMHC-I.- Since most nucleated cells in the body express class I MHC,most cells in the body can present antigen to cytotoxic Tcells. Although they are presenting antigen, these cells areusually not referred to as “antigen-presenting cells”. If theyare presenting antigen that will cause them to be killed bycytotoxic T cells, they are referred to as “target cells”.Professional vs Non-Professional APCs Remember: 1) MHC-II, 2) deliver co-stimulatorysignals Professional APC: DC MΦΦ B cells, why? DC: Always express high levels of MHC-IImolecules and co-stimulatory activity (B7molecule) MΦΦ: requires activation to up-regulate MHC-IImolecules and co-stimulatory molecules (B7molecules) B cells: always express MHC-II molecules butneeds to be activated to express co-stimulatoryactivity (B7 molecule)Self MHC Restriction Both MHC-I and MHC-II molecules canonly recognize antigens when presented bySELF-MHC molecules.ClassicExperiment toshow self -MHCrestrictionH-2KCD8 T cells No value for individual to have T cellsthat recognize foreign antigenassociated with foreign MHC Self MHC restriction occurs in thymusH-2K(-)H-2K( )H-2b(-)1

Ag processing is requiredRole for APC? Classical experiment showing that B and Tcells have different requirement for antigenrecognition. Processing is required for Th activation Processing is a metabolic active processPoints Concerning AntigenProcessing and Presentation1. Location of pathogen viruses in cytosol, MHC class I pathway, Tcresponse (Cytosolic pathway) extracellular bacteria, MHC class IIpathway, Th2 response Ab formation(Endocytic pathway) intracellular bacteria, MHC class IIpathway, Th1 response cellular response(Endocytic)Points Concerning AntigenProcessing and Presentation2. Peptides derived from both self andnon-self proteins can associate withMHC class I and class II molecules.3. Chemical nature of MHC groovedetermines which peptides it willbind.MHC-I and MHC-II associated with peptidesprocessed in different intracellularcompartmentsA) Class I MHC binds peptides derived from endogenousantigensB) Class I MHC binds peptides from antigens that havebeen processed via the cytosolic pathway (derivedfrom the cytoplasm of the cell)C) Class II MHC molecules bind peptides derived fromexogenous antigens. These antigens wereinternalized by phagocytosis or endocytosis.D) These peptides are said to have been processedwithin the endocytic pathway.2

Endogenous Pathway Peptides are generated by proteasomedegradation Peptides are transported from cytosol to theRER Peptides loading onto MHC-I is aided bychaperonesKuby Figure 8-5- The cytosolic antigen processing pathway - 2. The role of the TAP(Transporter associated with Antigenic Processing).- Size- HydrophobicityThe cytosolic antigen processing pathway - 3. Assembly of the class I-peptidecomplex1. The class I alpha chain isstabilized by calnexin.- Peptides from proteasome degradation of cytoplasmic proteins are transportedacross the membrane of the rough endoplasmic reticulum by a heterodimericprotein designated as TAP.2. When the alpha chainbinds beta-2-microglobulin:- calnexin is lost- calreticulin and tapasinbind- TAP is composed of two subunits - TAP1 and TAP-2- TAP-mediated transport is ATP-dependent3. Tapasin & calreticulinbrings the class Imolecules into the vicinityof the TAP.The genes for TAP1 and TAP2 are encoded within the MHC.4. A cytoplasmic peptidetransported through the TAPis loaded onto the class Imolecule.5. Class I MHC dissociatesfrom calreticulin and tapasin.Kuby Figure 8-6bClass I MHC-peptide complexis transported to Golgi and tothe cell surface.Kuby Figure 8-6aClass I MHC PathwayPeptide is presentedby MHC-I to CD8cytotoxic T cellPlasma membrane61 Viral protein is madeon cytoplasmicribosomesGlobular viralprotein - intact5Peptide passeswith MHC from Golgibody to surface2rERPeptide associateswith MHC-I complex4Golgi bodyClass II Processing:Peptide with MHCgoes to Golgi bodyMHC class I alphaand beta proteinsare made on the rERProteasomedegradesprotein topeptidesPeptide transporterproteinmovespeptide into ER3- The endocytic antigen processing pathway –processing of externally-derived peptides- Antigen can be taken into cells by various means:phagocytosis, endocytosis, pinocytosis, receptormediated endocytosis- Antigen taken up in these ways passes through aseries of intracellular compartments of increasingacidity - early endosome (pH 6.5-6.0), late endosome(pH 6.0-5.0), phagolysosome (pH 5.0)3

The endocytic antigen processingpathway - processing ofexternally-derived peptidesThree major events occur in theendosomal pathway:1) Degradation of material that wastaken in – endosome goes throughacidification and fusion withlysosome which contain a widearray of degradative enzymesThe endocytic antigen processingpathway - processing ofexternally-derived peptidesThe endosomal compartment iscompletely separate from theendoplasmic reticulum --- So,externally derived peptides areusually not loaded on to MHC-I.2) Loading of peptides from thismaterial on to class II MHCmolecules3) Transport of class II MHC - peptidecomplexes back to the cell surfaceFigure 5-18The endocytic antigen processingpathway - processing ofexternally-derived peptidesFigure 5-18Invariant Chain- guides transport toendocytic VesiclesClass II MHC is synthesized in the ER, butis not loaded with peptides therebecause it's peptide binding site isblocked by the invariant chain (Ii).Once class II MHC enters the endosomalcompartments, the invariant chain isdegraded, leaving a small fragment CLIP - in the peptide binding site.CLIP is removed by HLA-DM, which loadsa peptide on to class II MHCHLA-DO inhibits HLA-DM until the cell isactivatedCLIP Class II-associatedinvariant chain peptideHLA-DM – mediates exchangeHLA-DO – inhibits HLA-DMFigure 5-18Class II MHC PathwayPeptide MHC-IIcomplex is presentedto CD4 helper T cellReceptor (Ab)-Mediated EndocytosisGlobular proteinGlobular proteinCD4 helper T cell4Endosome fuses withplasma membraneImmunodominantpeptide bindsto class II MHCGolgibody3EndosomeFusion of endosomeand exocytic vesicleEndocytosis1Lysosome2Exocytic vesicle fuseswith endosomereleasing Ii from αβ dimerProtein is processed topeptides in endosomeor lysosomeClass II MHCSynthesisβ Ii 3 chains: α,β and IiEndoplasmic reticulumα4

Presentation of Non-PeptideAntigens CD1 molecules (CD1a-d)Structurally related to MHC-IEncoded outside the MHC regionPresent in APC (DC MØ B cells)Presents peptides of 12-22 aa in sizePresents to CD4, CD8 and NK cellsPresent LIPIDS and glycolipidsThe End!5

The role of the TAP (Transporter associated with Antigenic Processing). - Peptides from proteasome degradation of cytoplasmic proteins are transported across the membrane of the rough endoplasmic reticulum by a heterodimeric protein designated as TAP.-TAP is composed of two subunits - TAP1 and TAP-2 -