1 0/03/05; 9AM Some notes are provided here to help with the lecture on Antigen Presentation Protein Trafficking/The Secretory Pathway: From an immune perspective the secretory compartment and structures enclosed by vesicles are “seen” in different ways from proteins that reside in the cytosol or the nucleus. We will briefly review the secretory and endocytic pathways and discuss the biogenesis of membrane proteins. Some of the issues that will be discussed are summarized in Figures 1-3. Figure 1. An overview of the secretory pathway ERGolgiLysosomesEarly endosomesLateendosomesVesiculo-TubularCompartment Harvard-MIT Division of Health Sciences and TechnologyHST.176: Cellular and Molecular ImmunologyCourse Director: Dr. Shiv PillaiERGolgiMultivesicular andmultilamellar bodiesEarly endosomesLate endosomesProteasomes Figure 2. Protein degradation occurs mainly in lysosomes and proteasomes Proteins that enter the cell from the environment are primarily degraded in lysosomes. Most cytosolic and nuclear proteins are degraded in organelles called proteasomes. Intriguingly these two sites of degradation are each functionally linked to distinct antigen presentation pathways, different kinds of MHC molecules and the activation of different categories of T cells. Integral membrane proteins maybe inserted into the membrane in a number of ways, the two most common of these ways being considered in Figure 3.NCCNType IType IINCCNType IType IICell surfaceER or intracellularvesicles Figure 3. The orientation of Type I and Type II membrane proteins. Antigen presentation pathways Receptor mediated endocytosis will be discussed briefly in this lecture both in the context of the function of membrane bound immunoglobulins and in the context of antigen presentation pathways. T cells co-evolved with B cells. They have the ability to look “into” and destroy other host cells if the latter are infected. The basic design of T cell recognition depends on an intrinsic ability to ignore free or soluble antigens. T cell receptors can only recognize antigenic peptides that are appropriately exhibited on cell surfaces. The molecules that bring these peptides to the cell surface, and then exhibit them so that they may be scanned by passing T cells, are known as Major Histocompatibility Complex (MHC) proteins. These are the most polymorphic proteins known, encoded in most vertebrate species by a very large number of alleles. The antigen receptor on T cells is made up of two transmembrane polypeptides of the immmunoglobulin superfamily, the T Cell receptor (TCR)) α and β chains. Receptor diversity is generated by the rearrangement, in developing thymocytes, of gene segments that encode these polypeptides. The variable domains of the TCR α and β chains contribute to the antigen binding site. TCRs recognize cell surface MHC molecules which are complexed with peptides of either exogenous or endogenous origin. It is nowbelieved that all TCRs initially generated in the thymus are capable of recognizing MHC like shapes. We will return to this issue when we discuss why a relatively large proportion of T cells in any one individual have the potential to be alloreactive in later lectures on transplantation. The antigen recognizing TCR α and β chains associate with proteins of the CD3 complex, (CD3 γ, δ, ε, and ζ) which are required for the initiation of signaling from the antigen receptor. MHC proteins evolved at the same time as B and T cells, to provide a mechanism for T cell receptors to focus exclusively on antigenic fragments that are held on the cell surface. Since T cell receptors recognize only MHC-peptide complexes, free viruses or viral antigens are totally ignored by T lymphocytes. Although the physiological function of MHC molecules relates to antigen recognition by T cells, they acquired their name because they were discovered in the context of transplant rejection. They were originally referred to as Major Histocompatibility Complex Antigens because they were identified as the most prominent antigens involved in experimental graft rejection. Elegant genetic studies, which helped create the field of immunogenetics, established that these antigens map to a single chromosome in the mouse, to a set of genes that came to be known as the H-2 locus. The corresponding locus in man is on chromosome 6 and is referred to as the Human Leukocyte Antigen or HLA locus. MHC molecules in man are called HLA molecules and the terms MHC and HLA will be used interchangeably. HLA molecules are found on leukocytes but their expression is not restricted to white cells. MHC class I molecules are expressed on all nucleated cells and are recognized by T cell receptors on cytotoxic or CD8+ T cells (Cytotoxic T Lymphocytes or CTLs). They generally present peptides derived from proteins synthesized “endogenously” in the cell expressing the MHC class I molecule of interest. MHC class II molecules are primarily expressed on B cells and other professional antigen presenting cells (mainly macrophages and dendritic cells). Their expression may be induced on endothelial and epithelial cells exposed to cytokines such as γ-interferon. MHC class II molecules generally present peptides derived from exogenous proteins which must be internalized by the antigen presenting cell. These proteins are broken down or “processed” into peptides intracellularly. MHC class II-peptide complexes are recognized by antigen receptors on helper or CD4 + T cells. The structure of MHC proteins MHC molecules are cell surface glycoproteins which contain two membrane-proximal immunoglobulin like domains as well as two more distal specialized domains which together form a peptide binding groove embedded between two α-helical ridges. MHC class I molecules are made up of two polypeptide chains and a tightly associated peptide moiety (Figure 4). The MHC class I heavy chainis a transmembrane glycoprotein of about 40-45 kDa which associates tightly with a small non-anchored 12 kDa immunoglobulin superfamily protein known as β2 microglobulin. MHC class II molecules are heterodimers which consist of an α and a β chain, both of which are transmembrane glycoproteins (Figure 5). Our current understanding of MHC function owes a lot to the knowledge of the structure of MHC molecules obtained by X-ray crystallography. Crystal structures are available for a range of HLA class I and class II molecules complexed with specific peptides