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Antigen Receptors Study Guide

B Cell Receptors (BCRs)

• B cells rearrange their antibody gene segments as they mature
  • Each B cell generates one set of functional antibody genes (one VDJ gene for heavy chain and one VJ gene for light chain) as a result of this rearrangement
    • One VDJ gene is generated for heavy chain
    • One VJ gene is generated for light chain (either kappa or lambda)
  • Transcription of these genes allows formation of antibody molecules with a single combining site specificity that is able to interact with an antigenic determinant in a highly specific manner
• B cell receptors (BCRs) on their surface "display" the antibody combining site a mature B cell can make, thus allowing them to function in antibody responses
  • Secretion of antibodies by B cells results from their specific interaction with an antigenic determinant via their BCRs, which have two components:
    • membrane Ig (mIg) comprised of IgD and IgM monomers is coexpressed on the surface of naive (virgin) mature B cells
      • Variable domains containing three hypervariable (complementarity determining) regions comprise the N-terminal domains of both heavy and light chains
        • CDRs in variable domains of surface IgD and IgM bind antigenic determinants of native antigen
        • cross-linking of BCRs (capping) is thus initiated
      • Constant domains comprise the membrane-proximal domains of heavy chains (but not light chains, which do not interact with the B cell cytoplasmic membrane)
        • transmembrane segments (stretches of ~20 hydrophobic amino acids) near the C-terminal end of these heavy chains anchor them in B cell cytoplasmic membranes
        • cytoplasmic tails are nearly nonexistent at the C-terminal end of these heavy chains, so they are incapable of initiating signal transduction events, even though their N-terminal ends bind antigenic determinants
    • Ig-alpha/Ig-beta heterodimers, disulfide-linked polypeptides containing cytoplasmic domains with ITAMs (Immunoreceptor Tyrosine-based Activation Motifs), are associated with the mIg subunits (two heterodimer molecules per BCR)
      • heterodimers change conformation due to mIg cross-linking
      • ITAMs are then phosphorylated, generating a signal that is transmitted to the B cell nucleus via an intracellular signal transduction pathway
        • Ig-alpha has two ITAMs
        • Ig-beta has one ITAM
  • Clonal selection of B cells is based, then, upon the recruitment of individual B cells, via their ability to specifically interact with an antigenic determinant, into a proliferating population of cells that eventually differentiates into:
    • plasma cells, which actively produce and secrete antibody
    • B memory cells, which represent an expanded population of B cells able to respond to the inducing antigenic determinant

T Cell Receptors (TCRs)

• T cells rearrange their surface receptor gene segments as they mature
  • Receptor genes are generated as a result of this rearrangement
    • One VDJ gene is generated for beta chain
    • One VJ gene is generated for alpha chain
  • Transcription of these genes allows formation of receptor molecules with a single combining site specificity that is able to interact with a fragment of an antigenic molecule in a highly specific manner
• T cell receptors (TCRs) on their surface "display" the receptor combining site a mature T cell can make, thus allowing them to function in T cell-mediated responses (MHC-Ag-TCR interaction ... in motion)
  • TCRs are NOT secreted by T cells . . . instead . . .
  • T cell proliferation, differentiation and cytokine production result from their specific interaction with antigen fragments via their TCRs, which have two components:
    • TCR-alpha/TCR-beta heterodimers (in pairs)
      • Variable domains containing three hypervariable regions, also known as complementarity-determining regions (CDRs) comprise the N-terminal domains of both TCR-alpha and TCR-beta chains
        • CDRs in the variable domains of TCR-alpha and TCR-beta bind antigen fragments
        • antigen fragments must be bound to MHC molecules on antigen presenting cell (APC) surfaces to be recognized by TCR
      • Constant domains comprise the membrane-proximal domains of TCR-alpha and TCR-beta
        • transmembrane segments (stretches of ~20 hydrophobic amino acids) near the C-terminal end of the both constant domains anchor them to T cell cytoplasmic membranes
        • cytoplasmic tails are nearly nonexistent at the C-terminal ends of these polypeptides, so they are incapable of initiating signal transduction events, even though their N-terminal ends bind antigenic fragments
        • http://www.path.cam.ac.uk/~mrc7/functions/mhc_tcr1.html
    • CD3 must be coexpressed with the alpha/beta heterodimers
      • several polypeptides comprise CD3
        • gamma
        • delta
        • epsilon (in pairs)
        • zeta (these disulfide-linked dimers can be replaced by eta dimers, but zeta is the predominant molecule expressed in CD3)
      • charge-based (electrostatic) interactions link CD3 with the constant regions of alpha/beta herterodimers
      • Zeta and epsilon both have cytoplasmic domains with ITAMs (Immunoreceptor Tyrosine-based Activation Motifs)
        • zeta has three ITAMs (thus it initiates most of the signal transduction)
        • epsilon has one ITAM
      • TCR interaction with antigen fragments bound to MHC molecules on APCs initiates ITAM phosporylation by cytoplasmic protein kinases in T cells
      • ITAMs phosphorylation generates a molecular "signal" that is transmitted to the T cell nucleus via an intracellular signal transduction pathway
  • Clonal selection of T cells is based on recruitment of individual T cells, via their ability to specifically interact with an antigenic fragment, into proliferating populations of T cells that eventually differentiates into:
    • helper T (Th) cells, which actively produce and secrete cytokines
    • cytotoxic T cells (CTLs), which kill abnormal (virus-infected, tumor or graft) cells
    • T memory cells (either Th or CTL), which represent expanded populations of T cells able to respond to the inducing antigenic determinant

Antigen-Presenting Cell Receptors (ACRs)

• Antigen presenting cells (APCs) must process and present antigen before it can be recognized by TCRs
• Major Histocompatability Complex (MHC) molecules are the ACRs used by APCs for presentation of antigen fragments to T cells
  • MHC class I molecules
    • alpha chain has 3 domains plus a transmembrane and a cytoplasmic region
      • alpha 1 and alpha 2 domains are distal to the cell membrane and, together, they form the antigen binding site
      • alpha 3 domain has an Ig-fold conformation and is proximal to the cell membrane
      • transmembrane region is hydrophobic
      • cytoplasmic tail is hydrophilic
    • beta-2 microglobulin stabilizes the complex (in fact, it must be present for alpha chain to appear on the cell surface)
  • MHC class II molecules
    • alpha chain has 2 domains plus a transmembrane and a cytoplasmic region
      • alpha1 domain comprises half of the antigen-binding site
      • alpha2 domain has an Ig-fold conformation and is proximal to the cell membrane
      • alpha chain transmembrane region is hydrophobic and anchors the alpha chain to the cell membrane
      • alpha chain cytoplasmic tail is hydrophilic
    • beta chain has 2 domains plus a transmembrane and a cytoplasmic region
      • beta1 domain comprises the other half of the antigen-binding site
      • beta2 domain has an Ig-fold conformation and is proximal to the cell membrane
      • beta chain transmembrane region is hydrophobic and anchors the beta chain to the cell membrane
      • beta chain cytoplasmic tail is hydrophilic
• Genetics of the MHC
  • Early studies on the genetics of immune responses led to the discovery of the function of MHC molecules as genetically determined structures on immune cell surfaces that regulate immunological reactions
• Antigen fragments are generated by APCs and bound to MHC molecules during processing via one of two pathways:
  • Cytosolic antigen processing pathway generates antigen fragments that associate with class I MHC molecules on cell surfaces
    • proteins made within host cells (endogenous proteins) are tagged with ubiquitin, then degraded by proteasomes:
      • ATP-dependent complexes of peptidases comprised with multiple subunits
      • LMP2 and LMP7 (MHC-encoded Low Molecular weight Protein subunits) plus LMP10 (subunit that is not MHC-encoded) are found in proteasomes that generate peptides capable of binding to MHC-I molecules
    • antigen fragments generated by proteasomes are delivered to transporter proteins, TAP1 and TAP2 (Transporters associated with Antigen Processing) which are gatekeepers that facilitate antigen fragment entry into the endoplasmic reticulum, where they attach to the antigen binding sites of class I MHC molecules anchored there by interaction with TAP2, tapasin and calreticulin (all of which are proteins found on the inner leaf of the endoplasmic reticular membrane)
      • antigen fragment anchor position amino acid side chains, one at the C-terminus and another near the N-terminus, interact (via noncovalent bonding) with contact amino acid side chains in the alpha-helical and beta-pleated sheet regions of the alpha1 and alpha2 chain domains that comprise the antigen-binding groove
      • antigen fragments are generally 8-10 amino acids long, and fit tightly into the antigen-binding site:
        • fragments are anchored to MHC-I residues by interactions of their N-terminal (residues 1-3; nature varies) and C-terminal (8-9; hydrophobic or basic) amino acids
        • fragments longer than 8 amino acids bind at the ends, with the central region "poking" up out of the binding cleft like an "inchworm")
      • a wide variety of peptides can bind to each MHC I binding site, provided they can fit in the antigen-binding groove and possess the appropriate amino acids to anchor the ends
      • LMP2, LMP7 and LMP10 appear to foster generation of fragments 8-10 amino acids in length, with hydrophobic or basic residues at the C-terminus (due to protease specificity of these subunits)
    • antigen fragment binding facilitates release of MHC I molecules from their association with TAP2 so they can traverse the Golgi system and be transported to the cytoplasmic membrane in vesicles, which fuse with the cytoplasmic membrane
    • fusion of vesicles allows MHC I molecules to be anchored in, and located on the outer side of, the cytoplasmic membrane
    • MHC class I molecules with bound antigen fragments (linear peptide samples of all the proteins being made by the cell) are thus displayed on cytoplasmic membranes of all cells (view another version ... view it in 3D)
  • Endosomal antigen processing pathway generates antigen fragments that associate with class II MHC molecules on cell surfaces (view another version of this pathway)
    • extracellular proteins that enter cells by endocytosis (pinocytosis or phagocytosis) are degraded by lysosomal enzymes in early endosomes
    • late endosomes fuse with transport vesicles containing MHC II molecules whose antigen binding sites are filled with invariant chain
    • invariant chain, except for a small peptide called class II-associated invariant chain peptide or CLIP) is digested by the lysosomal enzymes
    • antigen fragments bind to MHC II antigen-binding sites as HLA-DM catalyzes removal of CLIP
      • antigen fragment anchor position amino acid side chains along the entire length of the antigen fragment interact (via noncovalent bonding) with contact amino acid side chains in the alpha-helical and beta-pleated sheet regions of the antigen-binding groove comprised by the alpha1 and beta1 chain domains
      • antigen fragments are generally 10-34 amino acids long, with about 9 amino acids actually binding in the antigen-binding cleft and the rest hanging out each end of the groove (binding may actually occur prior to final cleavage of the peptide fragment to its 9-34 residue length)
      • a wide variety of peptides can bind to each MHC II binding site
    • late endosome is transported to, and fuses with, the cytoplasmic membrane, with MHC II molecules anchored on the outside
    • MHC class II molecules with bound antigen fragments (linear peptide samples of all the proteins being made outside the cell) are thus displayed on cytoplasmic membranes of APCs