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Processing of intracellular antigen for presentation by class I MHC


Processing of intracellular antigen for presentation by class I MHC
Proteasomes are constitutively involved in the routine turnover and cellular degradation of proteins. Cytosolic proteins destined for antigen presentation, including viral proteins, are degraded to peptides via the pathway involving these structures. In addition to proteins that are already present in the cytosol, misfolded and misassembled proteins are transported from the ER back into the cytosol by a quality‐control process referred to as ER‐associated protein degradation (ERAD). Proteins that have undergone retrotranslocation from the ER into the cytosol can then also be processed for class I presentation, as can proteins derived from mitochondria. Prior to processing, polypeptide antigens are covalently linked to several molecules of the 7.5 kDa protein ubiquitin in an ATP‐dependent process. This polyubiquitination targets the polypeptides to the proteasome (Figure 5.15).


Figure 5.15 Processing of endogenous antigen and presentation by class I MHC. Cytosolic proteins (a) targeted for degradation become polyubiquitinated by the addition of several molecules of ubiquitin. The ubiquitinated protein binds to the 19S regulator of the proteasome (b) which in a ATP‐dependent reaction removes the ubiquitin, unfolds the protein, and pushes it into the cylindrical structure of the 20S core proteasome that is made up of 28 subunits arranged in four stacked rings. The resulting peptides are transported into the endoplasmic reticulum (ER) by TAP1 and TAP2 (c). Under the influence of the peptide loading complex (PLC; which comprises TAP1/2 together with calreticulin, tapasin, and ERp57) the peptides are loaded into the groove of the membrane‐bound class I MHC. ERp57 isomerizes disulfide bonds to ensure the correct conformation of the class I molecule. Tapasin forms a bridge between TAP1/2 and the other PLC components and is covalently linked to ERp57, which in turn is noncovalently bound to the calreticulin. Following peptide loading the peptide–MHC complex is released from the PLC (d), traverses the Golgi system (e), and appears on the cell surface (f) ready for presentation to the T‐cell receptor. Mutant cells deficient in TAP1/2 do not deliver peptides to class I and cannot function as cytotoxic T‐cell targets.

Only a small minority of the peptides produced by the housekeeping proteasome are the optimal length (8–10 amino acids) to fit into the MHC class I groove; the remainder are either too short or too long. Longer peptides can be subjected to additional processing by, for example, cytosolic aminopeptidases (such as leucine aminopeptidase). Processing can also occur following transfer into the ER; in humans using the endoplasmic reticulum resident aminopeptidases (ERAP1 and ERAP2), and in mice the ER aminopeptidase associated with antigen processing (ERAAP). If the peptides are only slightly too long they can still bind to the groove and be recognized by TCRs but in this case, which may apply to up to 10% of class I‐bound peptides, they bulge out from the groove.
The cytokine IFNγ increases the production of three specialized catalytic proteosomal subunits, β1i, β2i, and β5i, which replace the homologous catalytic subunits in the housekeeping proteasome to produce the immunoproteasome, a structure with modified cleavage specificity that greatly increases the proportion of 8–10 amino acid long peptides generated. Both proteasome and immunoproteasome generated peptides are translocated into the ER by the heterodimeric transporter associated with antigen processing (made up of TAP1 and TAP2 subunits) (Figure 5.15). The newly synthesized class I α chain is retained in the ER by the lectin‐like chaperone calnexin, which binds to the monoglucosylated N‐linked glycan of the nascent α chain. Calnexin assists in protein folding and promotes assembly with β2‐microglobulin. The calnexin is then replaced with calreticulin, which has similar lectin‐like properties and, together with TAP1/2, tapasin, and ERp57 (57 kDa ER thiol oxidoreductase), constitutes the peptide loading complex (PLC). The tapasin bridge ensures that the empty class I molecule sits adjacent to the TAP pores in the ER, thereby facilitating the loading of peptides. Tapasin also plays a peptide‐editing role, ensuring preferential incorporation of peptides with high‐affinity binding to the MHC class I molecules. Upon peptide loading, the class I molecule dissociates from the PLC, and the now stable MHC–peptide traverses the Golgi stack and reaches the surface where it is a sitting target for the cytotoxic T‐cell.