8536d_ch08_185-199 8/2/02 10: 08 AM Page 193 mac79 Mac 79: 45_ Bwppldsby et al./ Immunology 5e 193 B2 microglobulin → Exit RER 9 Class I mHC Calnexin CaInexin-associated Calreticulin-tapasin Class I mhc class I mHc a chain associated class I molecule MHC molecule Tapasin Calreticulin Tapasin Calreticulin IGURE 8-7 Assembly and stabilization of class I MHC mole- chaperonin calreticulin and to tapasin, which is associated with the cules. Newly formed class I a chains associate with calnexin, a peptide transporter TAP. This association promotes binding of an molecular chaperone, in the RER membrane. Subsequent binding antigenic peptide, which stabilizes the class I molecule-peptide to B2-microglobulin releases calnexin and allows binding to the complex, allowing its release from the reR heterodimer with the TAP protein(see Figure 8-6b)pro- the experiment shown in Figure 8-3 demonstrated, internal- motes peptide capture by the class I molecule before the pep- ized antigen takes 1-3 h to transverse the endocytic pathway tides are exposed to the luminal environment of the RER. and appear at the cell surface in the form of peptide-class Il Peptides not bound by class I molecules are rapidly degraded. MHC complexes. The ende ay appears to involve As a consequence of peptide binding, the class I molecule dis- three increasingly acidic compartments: early endosomes (pH plays increased stability and can dissociate from calreticulin 6.0-6.5); late endosomes, or endolysosomes(pH5.0-6.0); and and tapasin, exit from the RER, and proceed to the cell sur- lysosomes(pH 4.5-5.0). Internalized antigen moves from face via the Golgi. An additional chaperone protein, ERp57, early to late endosomes and finally to lysosomes, encountering has been observed in association with calnexin and calretic- hydrolytic enzymes and a lower pH in each compartment(Fig ulin complexes. The precise role of this resident endoplasmic ure 8-9). Lysosomes, for example, contain a unique collection reticulum protein in the class I peptide assembly and loading of more than 40 acid-dependent hydrolases, including pro- process has not yet been defined, but it is thought to con- teases, nucleases, glycosidases, lipases, phospholipases, and tribute to the formation of disulfide bonds during the matu- phosphatases. Within the compartments of the endocytic tion of class I chains. Because its role is not clearly defined, pathway, antigen is degraded into oligopeptides of about 13- ERp57 is not shown in Figures 8-6 and 8-7. 18 residues, which bind to class lI MHC molecules. Because the hydrolytic enzymes are optimally active under acidic condi tions (low pH), antigen processing can be inhibited by chem Exogenous Antigens: The Endocytic cal agents that increase the pH of the compartments (e.g chloroquine)as well as by protease inhibitors(e. g, leupeptin Pathway The mechanism by which internalized antigen moves Figure 8-8 recapitulates the endogenous pathway discussed from one endocytic compartment to the next has not bee previously (left side), and compares it with the separate exoge conclusively demonstrated. It has been suggested that early nous pathway (right), which we shall now consider Whether ndosomes from the periphery move inward to become late an antigenic peptide associates with class I or with class ll mol ndosomes and finally lysosomes. Alternatively, small trans- port vesicles may carry antigens from one compartment to ecules is dictated by the mode of entry into the cell, either ex- the next. Eventually the endocytic compartments,or por- ogenous or endogenous, and by the site of processing Antigen-presenting cells can internalize antigen by phago- ns of them, return to the cell periphery, where they fuse cytosis, endocytosis, or both. Macrophages internalize antigen th the plasma membrane. In this way, the surface receptor by both processes, whereas most other APCs are not phago- re recycled. cytic or are poorly phagocytic and therefore internalize exoge- nous antigen only by endocytosis(either receptor-mediated The Invariant Chain Guides Transport endocytosis or pinocytosis). B cells, for example, internalize of Class II MHC Molecules ntigen very effectively by receptor-mediated endocytosis u to Endocytic Vesicles ing antigen-specific membrane antibody as the receptor. Since antigen-presenting cells express both class I and class Il Peptides Are Generated from Internalized MHC molecules, some mechanism must exist to prevent Molecules in Endocytic Vesicle lass II MHC molecules from binding to the same set of anti- nic peptides as the class I molecules. When class II MHC Once an antigen is internalized, it is degraded into peptides molecule are synthesized within the RER, three pairs of class vithin compartments of the endocytic processing pathway. As II aB chains associate with a preassembled trimer of aAntigen Processing and Presentation CHAPTER 8 193 heterodimer with the TAP protein (see Figure 8-6b) pro￾motes peptide capture by the class I molecule before the pep￾tides are exposed to the luminal environment of the RER. Peptides not bound by class I molecules are rapidly degraded. As a consequence of peptide binding, the class I molecule dis￾plays increased stability and can dissociate from calreticulin and tapasin, exit from the RER, and proceed to the cell sur￾face via the Golgi. An additional chaperone protein, ERp57, has been observed in association with calnexin and calretic￾ulin complexes. The precise role of this resident endoplasmic reticulum protein in the class I peptide assembly and loading process has not yet been defined, but it is thought to con￾tribute to the formation of disulfide bonds during the matu￾ration of class I chains. Because its role is not clearly defined, ERp57 is not shown in Figures 8-6 and 8-7. Exogenous Antigens: The Endocytic Pathway Figure 8-8 recapitulates the endogenous pathway discussed previously (left side), and compares it with the separate exoge￾nous pathway (right), which we shall now consider. Whether an antigenic peptide associates with class I or with class II mol￾ecules is dictated by the mode of entry into the cell, either ex￾ogenous or endogenous, and by the site of processing. Antigen-presenting cells can internalize antigen by phago￾cytosis, endocytosis, or both. Macrophages internalize antigen by both processes, whereas most other APCs are not phago￾cytic or are poorly phagocytic and therefore internalize exoge￾nous antigen only by endocytosis (either receptor-mediated endocytosis or pinocytosis). B cells, for example, internalize antigen very effectively by receptor-mediated endocytosis us￾ing antigen-specific membrane antibody as the receptor. Peptides Are Generated from Internalized Molecules in Endocytic Vesicles Once an antigen is internalized, it is degraded into peptides within compartments of the endocytic processing pathway. As the experiment shown in Figure 8-3 demonstrated, internal￾ized antigen takes 1–3 h to transverse the endocytic pathway and appear at the cell surface in the form of peptide–class II MHC complexes. The endocytic pathway appears to involve three increasingly acidic compartments: early endosomes (pH 6.0–6.5); late endosomes, or endolysosomes (pH 5.0–6.0); and lysosomes (pH 4.5–5.0). Internalized antigen moves from early to late endosomes and finally to lysosomes, encountering hydrolytic enzymes and a lower pH in each compartment (Fig￾ure 8-9). Lysosomes, for example, contain a unique collection of more than 40 acid-dependent hydrolases, including pro￾teases, nucleases, glycosidases, lipases, phospholipases, and phosphatases. Within the compartments of the endocytic pathway, antigen is degraded into oligopeptides of about 13– 18 residues,which bind to class II MHC molecules.Because the hydrolytic enzymes are optimally active under acidic condi￾tions (low pH), antigen processing can be inhibited by chemi￾cal agents that increase the pH of the compartments (e.g., chloroquine) as well as by protease inhibitors (e.g., leupeptin). The mechanism by which internalized antigen moves from one endocytic compartment to the next has not been conclusively demonstrated. It has been suggested that early endosomes from the periphery move inward to become late endosomes and finally lysosomes. Alternatively, small trans￾port vesicles may carry antigens from one compartment to the next. Eventually the endocytic compartments, or por￾tions of them, return to the cell periphery, where they fuse with the plasma membrane. In this way, the surface receptors are recycled. The Invariant Chain Guides Transport of Class II MHC Molecules to Endocytic Vesicles Since antigen-presenting cells express both class I and class II MHC molecules, some mechanism must exist to prevent class II MHC molecules from binding to the same set of anti￾genic peptides as the class I molecules. When class II MHC molecule are synthesized within the RER, three pairs of class II  chains associate with a preassembled trimer of a FIGURE 8-7 Assembly and stabilization of class I MHC mole￾cules. Newly formed class I  chains associate with calnexin, a molecular chaperone, in the RER membrane. Subsequent binding to 2-microglobulin releases calnexin and allows binding to the chaperonin calreticulin and to tapasin, which is associated with the peptide transporter TAP. This association promotes binding of an antigenic peptide, which stabilizes the class I molecule–peptide complex, allowing its release from the RER. + + Peptides Exit RER Calnexin Calnexin Class I MHC α chain Class I MHC molecule Calreticulin-tapasin– associated class I MHC molecule Calnexin-associated class I MHC α chain β2 microglobulin + + Tapasin Calreticulin Tapasin Calreticulin 8536d_ch08_185-199 8/2/02 10:08 AM Page 193 mac79 Mac 79:45_BW:Goldsby et al. / Immunology 5e: