Antigen Processing

Antigen Processing and Presentation, MHC Restriction and Role of Thymus I.

Comparison of BCR and TCR

B cells and T cells recognize different substances as antigens and in a different form. The B cell uses cell surface-bound immunoglobulin as a receptor and the specificity of that receptor is the same as the immunoglobulin that it is able to secrete after activation. B cells recognize the foll follow owin ing g anti antige gens ns in solu solubl ble e form form:: 1) prot protei eins ns (bot (both h conf confor orma mati tion onal al dete determ rmin inan ants ts and and determinants exposed by denaturation or proteolysis); 2) nucleic acids; 3) polysaccharides; 4) some lipids; 5) small chemicals (haptens). In contrast, the overwhelming majority of antigens for T cells are proteins, and these must be fragm fragment ented ed and recogn recognize ized d in associ associati ation on with with MHC MHC produc products ts expres expressed sed on the surfac surface e of  nucleated cells, not in soluble form. T cells are grouped functionally according to the class of  MHC molecules that associate with the peptide fragments of protein: helper T cells recognize only those peptides associated with class II MHC molecules, and cytotoxic T cells recognize only those peptides associated with class I MHC molecules. II. II.

Anti Antig gen Pr Proces ocess sing ing and and Prese esentati tation on

Antig Antigen en proces processin sing g refer refers s to the ability ability of and APCs to break break down down a protei protein n antige antigen n into into peptides peptides and to associate associate those those peptides peptides with MHC molecule molecules. s. Antigen Antigen presentat presentation ion is the process of displaying peptide antigens associated with MHC molecules to a T cell. The path leading to the association of protein fragments with MHC molecules differs for class I and class II MHC. MHC. MHC MHC clas class s I mole molecu cule les s pres presen entt degr degrad adat atio ion n prod produc ucts ts deri derive ved d from from intr intrac acel ellu lula larr (endogenous) proteins in the cytosol. MHC class II molecules present fragments derived from extracellular (exogenous) (exogenous) proteins that are located in an intracellular intracellular compartment. A.

Class Class I MHC Pathw Pathway ay - All All nucleate nucleated d cells express express class class I MHC. MHC. As shown shown in Figure Figure 1, proteins proteins are fragmented in the cytosol by proteosomes (a complex of proteins having proteolytic activity) or by other proteases. The fragments are then transported across the membrane of  the endoplasmic reticulum by transporter

Figure 1. Antigen Processing and Presentation - Class I MHC Pathway

Antigen Antigen Processing Processing and Presentation Presentation - Class I MHC Pathway proteins. (The transporter proteins and some components of the proteosome are encoded by genes in the MHC complex). Synthesis and assembly of class I heavy chain and β2- microglobulin occurs in the endoplasmic reticulum. Within the endoplasmic reticulum, the MHC class I heavy chain, β2-microglobulin and peptide form a stable complex that is transported to the cell surface.

B.

Class II MHC pathway - Whereas all nucleated cells express class I MHC, only a limited group of cells express class II MHC, which includes the antigen presenting cells (APC). The principal APC are macrophages, dendritic cells (Langerhans cells), and B cells, and the expres expressio sion n of class class II MHC MHC molec molecule ules s is either either consti constitut tutive ive or induc inducibl ible, e, especi especiall ally y by interferon-gamma interferon-gamma in the case of macrophages. macrophages. As shown in Figure Figure 2, exogenous proteins proteins taken in by endocytosis are fragmented by proteases in an endosome. The alpha and beta chains of MHC class II, along with an invariant chain, are synthesized and assembled in the endoplasmic endoplasmic reticulum. The invariant chain prevents prevents endogenous peptides peptides from the cytosol from from associa associatin ting g with with class class II MHC MHC molec molecule ules. s. The class class II MHC MHC molecu molecules les with with the associated invariant chain are transported through the Golgi and trans-Golgi apparatus to reach the endosome, where the invariant chain is digested, and the peptide fragments from the exogenous protein are able to associate with the class II MHC molecules, which are finally transported transported to the cell surface.

Figure 2. Antigen Processing and Presentation - Class II MHC Pathway C. Other Other points points conc concern erning ing antig antigen en proce process ssing ing and and present presentati ation on 1. One way of rationaliz rationalizing ing why two differen differentt antigen process processing ing pathways pathways developed developed is to say that each ultimately stimulates the population of T cells that is most effective in eliminating a particular type of antigen. Viruses replicate within nucleated cells in the cytosol and produce

endogenous antigens that can associate with class I MHC. By killing these infected cells, cytotoxic T cells help to control the spread of the virus. Bacteria mainly reside and replicate extracellularly. By being taken up and fragmented inside cells as exogenous antigens that can associate with class II MHC molecules, helper Th2 T cells can be activated to assist B cells to make make antibo antibody dy agains againstt bacter bacteria, ia, which which lim limits its the growth growth of these these organ organism isms. s. Some Some bacteria grow intracellularly inside the vesicles of cells like macrophages. macrophages. Inflammatory Th1 T cells help to activate macrophages to kill the intracellular bacteria. 2. Fragment Fragments s of self, as well as non-self, non-self, proteins proteins associat associate e with MHC molecul molecules es of both classes and are express expressed ed at the the cell cell surfac surface. e. T cells cells capable capable of recogn recognizi izing ng self peptide peptides s are eliminated during development (Clonal Selection Postulate) 3. Which Which protein protein fragme fragments nts bind is a funct function ion of the chemica chemicall nature nature of the groove groove for that that specific MHC molecule I. Self MHC Restriction

In order for a T cell to recognize and respond to a foreign protein antigen, it must recognize the MHC on the presenting presenting cell as self MHC. This is term termed ed self self MHC MHC rest restri rict ctio ion. n. Help Helper er T cell cells s reco recogn gniz ize e anti antige gen n in cont contex extt of clas class s II self  self  MHC. MHC. Cytoto Cytotoxic xic T cells cells recogn recognize ize antige antigen n in cont contex extt of clas class s I self self MHC. MHC. The The proc proces ess s whereby T cells become restricted to recognizing self MHC molecules occurs in the thymus. thymus. The experi experiment mental al basis basis of self self MHC restriction for APC-helper T cell interactions and for class I MHC-cytotoxic T cell interactions are shown in Figures 3 and 4, respectively. respectively.

Figu Figure re 3. Self Self MHC MHC Restr Restric icti tion on of Th/AP Th/APC C Interactions.

Figure 4. Self MHC Restriction of Tc/ target cell Interactions

II.

Antigen Pr Presenting Ce Cells

 The three main types of antigen presenting cells are dendritic cells, macrophages and B cells, although other cells, that express class II MHC molecules, (e.g., thymic epithelial cells) can act as antig antigen en present presentin ing g cells cells in some some cases cases.. Dendri Dendritic tic cells, cells, which which are are found found in skin skin and and other other tissues, ingest antigens by pinocytosis and transport antigens to the lymph nodes nodes and spleen. spleen. In the lymph lymph nodes and spleen they are found predominantly in the T cells areas. areas. Dendritic Dendritic cells cells are the most most effective effective antigen presentin presenting g cells and can present present antig antigen ens s to naïve naïve (virg (virgin) in) T cells. cells. Furthe Furtherm rmor ore, e, they they can can presen presentt intern internali alized zed antige antigens ns in associa association tion with either class I or class class II MHC molecules molecules (cross (cross presentat presentation), ion), although although the predo predomin minan antt pathw pathway ay for interna internaliz lized ed antig antigen en is the class class II pathwa pathway. y. The The secon second d type type of  antig antigen en prese presenti nting ng cell cell is the the macro macropha phage. ge. These These cells ingest ingest antige antigen n by phagocy phagocytos tosis is of  pinocytosis. pinocytosis. Macrophages Macrophages are not as effective in presenting antigen to naïve T cells but they are very good in activating activating memory memory T cells. The third type type of antigen presenting presenting cell is the B cell.   The These se cell cells s bind bind anti antige gen n via via thei theirr surf surfac ace e Ig and and inge ingest st anti antige gens ns by pino pinocy cyto tosi sis. s. Like Like macrophages these cells are not as effective as dendrite cells in presenting antigen to naïve T cells. cells. B cells are very effective effective in presenting presenting antigen antigen to memory memory T cells, cells, especially especially when the antigen concentration concentration is low because surface Ig on the B cells binds antigen with a high affinity. III.

Presentation of Superantigens Superantigens

Superantigens are antigens that can polyclonally activate T cells (see lecture on antigens) to produce large quantities quantities of cytokines that can have pathological pathological effects. These antigens must be presented to T cells in association with class II MHC molecules but the antigen does not need to be processed. Figure 5 compares compares how conventional conventional antigens and superantigens superantigens are presented presented to T cells. In the case of a superantigen superantigen the intact protein binds to class class II MHC molecules and and to one or more more Vβ regions of the the TCR. The antigen is is not bound to the peptide binding groove groove of  the MHC MHC molecul molecule e or to the antigen antigen bindi binding ng site of the TCR. Thus, Thus, any T cell that uses uses a particular Vβ in its TCR will be activated activated by a superantigen, superantigen, resulting in the activation activation of a large numbers of T cells. cells. Each superantigen superantigen will bind to a different set set of Vβ regions.

IV.

Role of Thymus

Both Th and Tc cells are self-MHC restricted. In healthy, normal addition, T cells do not normally recognize selfantigens antigens.. How are are self MHC restr restricted icted T cells cells generated and why are self-reacting T cells not produ produced ced? ? Rando Random m VDJ rearr rearrang angeme ements nts in T cells would be expected to generate some T cells that can recognize non-self MHC and some   T cells cells that can can recognize recognize self-a self-antige ntigens. ns. It is the role of the thymus to ensure that the only T cells cells that that get to the perip peripher hery y are are self-M self-MHC HC restricted and unable to react with self-antigen. Func Functi tion onal al T cell cells s in the the peri periph pher ery y have have to recognize foreign antigens associated with self  MHC, MHC, beca becaus use e AP APC C or targ target et cell cells s pres presen entt fore foreig ign n anti antige gen n asso associ ciat ated ed with with self self MHC. MHC. However ever,, an individual does not need functional T cell ells in the periphery that recognize recognize antigen antigen (self (self or foreign) foreign) associa associated ted with foreign MHC. An individual individual especially does not want functional T cells in the periphery that Figure 5. Presentation of Superantigens can recognize self-antigens associated with self  MHC MHC becaus because e they they could could lead lead to damage damage of  As a resu result lt of rand random om VD VDJJ reco recomb mbin inat atio ion n events occurring in immature T cells within the thymus, TCRs of all specificities are produced. Processes in the thymus determine which TCR spec specif ific icit itie ies s are are reta retain ined ed.. Ther There e are are two two sequenti sequential al steps steps shown shown in Figure Figure 6. 6. First, First, T cell cells s with with the the abil abilit ity y to bind bind to self self MHC molec olecul ules es expr expres esse sed d by cort cortic ical al thym thymic ic epithelial cells are are retained. This is known as as posit positive ive selec selectio tion. n. Those Those that do not not bind, undergo apoptosis. apoptosis. Thus, T cells having having a TCR that recognizes self MHC survive. Next, T cells with the ability to bind to self MHC molecules associa associated ted with self-mo self-molecul lecules es expresse expressed d by thym thymic ic epit epithe heli lial al cell cells, s, dend dendri riti tic c cell cells s and and macr macrop opha hage ges s are are kill killed ed.. This This is know known n as negative selection. Those that do not bind are retain retained. ed. As a resul resultt of these these two steps steps,, T cells having a TCR that recognizes self MHC and foreign foreign antigen antigen survi survive. ve. Each T cell cell that survives positive and negative selection in the thym thymus us and and is rele releas ased ed into into the the peri periph pher ery y retains its specific T cell receptor (TCR). While positive and negative selection is occurring in the the thym thymus us the the im imma matu ture re T cell cells s are are also also expr expres essi sing ng CD4 CD4 or CD8 CD8 anti antige gens ns on thei theirr

tissues.

surface. Initially the the pre T cell that enters the the thym hymus is CD4CD4-C CD8 D8--. In the the thymus it beco become mes s CD4+ CD4+CD CD8+ 8+ and and as posi positi tive ve and and negati negative ve select selection ion procee proceeds ds a cell cell become becomes s either a CD4+ CD4+ or CD8+ CD8+ cell. The commitment commitment to becom ecome e eith ither a CD4 CD4+ or CD8+ cells ells depends on which class of MHC molecule the cell cell enco encoun unte ters rs.. If a CD4+ CD4+CD CD8+ 8+ cell cell is presented with a class I molecule it will down regulate CD4 and become a CD8+ cell. If a cell is presented with a class II MHC molecule it will down regulate CD8 and become a CD4+

cell

Figure Figure 6. Positiv Positive e and negative negative selectio selection n in the thymus.

Figure 7. Generation of CD4+ or CD8+ cells in the thymus. V.

Nega egativ tive se selecti ection on in the the pe perip riphery hery

Positive and negative negative selection in the thymus is not a 100% efficient process. process. In addition, not all self-antigens self-antigens may be expressed expressed in the thymus. thymus. Thus some some self-reactive T cells cells may get to the periphery. Thus, there are additional mechanisms mechanisms that are designed to eliminate eliminate self-reactive T cells in the periphery periphery.. These These will be discusse discussed d in the tolerance tolerance lecture. lecture. Since Since B cells cells are not MHC-restricted MHC-restricted there is no need for positive selection of B cells. However, negative selection (i.e., elimination of self-reactive self-reactive clones) of of B cells is required. This occurs during during B cell development in the bone marrow. However, negative negative selection of B cells is not a critical as for T cells since, in most instances, instances, B cells require T cell help in order order to become activated. Thus, if a self-reactive self-reactive B cell does get to the periphery it will not be activated due to lack of T cell help.