Receptors and Signaling: B- and T-Cell Receptors

Essential Concepts

  • A cellular signal instructs a cell to change its metabolic or proliferative state.
  • Signals are generated by the binding of a ligand to a complementary cell-bound receptor.
    • Ligand: A molecule that binds to another molecule.
    • Cognate: Describes two biomolecules that normally interact (e.g., an enzyme and its normal substrate, a receptor and its normal ligand).
  • A cell's susceptibility to a ligand's actions can be altered by modulating the expression of the receptor for that ligand.
  • The ligand can be a soluble molecule or a peptide, carbohydrate, or lipid on a cell surface.
  • The ligand may travel long distances in the bloodstream or lymphatics to reach a cell with the cognate receptor.

Receptor-Ligand Interactions

  • Receptor-ligand binding occurs via multiple noncovalent bonds.
    • Each individual bond may be weak, but the cumulative effect of many bonds provides great strength.
  • Ligand-receptor binding induces a molecular change in the receptor:
    • Conformational change.
    • Dimerization.
    • Covalent modification.
  • Receptor alterations induce cascades of intracellular events:
    • Activation of enzymes.
    • Changes in intracellular locations of molecules.

Receptor Conformational Change

  • A signaling molecule (ligand) binds to a ligand-gated ion channel receptor.
  • This binding induces a conformational change, opening a gate and allowing ions to flow across the plasma membrane.
  • This leads to a cellular response.

Ligand-Induced Receptor Dimerization

  • Antigen-mediated receptor dimerization is a common result of ligand binding.
  • Multimerization can also occur, where more than two receptors come together.

Ligand-Induced Receptor Covalent Modification (Phosphorylation)

  • A signal molecule binds to inactive receptor tyrosine kinases (RTKs).
  • This leads to cross-phosphorylation by activated kinase domains in the cytosol.
  • Covalent modification involves the attachment of a small chemical group.

Receptor-Ligand Interactions Enhanced by Co-receptor Binding

  • Antigen-immune system receptor interactions are often enhanced by co-receptor binding.
  • Co-receptor interactions are separate receptor-ligand interactions that occur near the original interaction.
  • A single type of interaction may be insufficient for activation, and a co-receptor interaction provides a second signaling interaction to further signal the cell to proceed with activation.

CD8 Co-receptor Example

  • An infected cell presents viral peptides on Class I histocompatibility molecules.
  • The CD8 co-receptor on a cytotoxic T cell interacts with the Class I molecule, while the T cell receptor interacts with the viral peptide.

Multivalent Ligands

  • A multivalent ligand is present as multiple copies on the cell surface, allowing the simultaneous binding of multiple receptors.

Receptor-Ligand Interactions: Affinity vs. Avidity

  • Receptor-antigen interactions are usually multivalent, increasing avidity.
  • Affinity is the strength of an individual pairing between a receptor and ligand.
  • Avidity is the combined strength of multiple interactions.
  • An interaction may have weak affinity but high overall avidity.

Receptor and Ligand Expression

  • Receptor and ligand expression can vary during an immune response.
  • Example: White blood cells treated with an activating mitogen upregulate the receptor for cytokine IL-2.

Local Concentrations of Cytokines

  • Local concentrations of cytokines and other ligands can be extremely high.
  • Cells may direct their secretion machinery toward a recipient for maximum effect (vectorial redistribution of secretory apparatus).
  • Example: Dendritic cells secrete cytokine IL-12 to T cells, with the cytokine-filled area localized in the dendritic cell.

Signaling Pathways

  • Binding of antigen to receptor induces an internal signaling cascade.
  • This cascade leads to cellular alterations in:
    • Motility.
    • Adhesive properties.
    • Transcriptional programming.
  • These cascades drive cellular changes during an immune response against an antigen.
  • The same proteins are often used in different cell types, but the triggering receptors may differ.

Signal Transduction

  • The process involves receptor binding, signal transduction, and a cellular response.
  • Key components include receptor-associated molecules, ligands, receptors, co-receptors, receptor-associated tyrosine kinases, PIP2, PIP3, DAG, PLCγ, PI3 kinase, IP3, adapter proteins (SOS, Ras-GRP), Ras, and various downstream targets.
  • This leads to Ca2+ release, calmodulin and calcineurin activation, phosphorylation, ubiquitination, activation of MAP kinase cascade, NFAT, AP-1, and NF-κB, ultimately resulting in gene activation.

Receptors Requiring Receptor-Associated Molecules

  • Some receptors require receptor-associated molecules to signal cell activation.
  • B- and T-cell receptors have short cytoplasmic portions.
  • Other molecules associate with them for signal transduction.
    • B cells use Igα and Igβ molecules.
    • T cells use CD3 complexes and CD28 molecules.

B-Cell Receptor (BCR)

  • The antigen receptor on B cells is a membrane-bound form of immunoglobulin.

The B-Cell Receptor (BCR) Complex

  • All isotypes of mIg have short cytoplasmic tails.
    • mIgM and mIgD: three amino acids long.
    • mIgA: 14 amino acids long.
    • mIgG and mIgE: 28 amino acids long.
  • These tails are too short to interact with intracellular signaling machinery.
  • mIg interacts with a disulfide-linked heterodimer called Ig-α/Ig-β, forming the BCR complex.

Antibody Structure

  • Immunoglobulin = antibody.
  • Two heavy chains (variable and constant regions).
  • Two light chains (variable and constant regions).
  • Held together by intra-/interchain disulfide covalent bonds.
  • Secreted antibodies are grouped into five major classes, differentiated by the amino acid sequence of the heavy chain constant region.
  • Each class performs different functions during immune responses.
  • Except for their variable regions, all immunoglobulins within one class share approximately 90% homology, but only 60% among the other classes.

Antibody Domains and Functions

  • CH1 and CL domains extend the “arms” of the antibody away from the hinge region.
  • VH and VL domains, located at the “top of the Y,” enable the Ab molecule to bind to its specific antigen.
  • Each Ab can bind two antigen molecules.

Fragment Crystallizable Region (Fc Region)

  • The Fc region is the tail region of an antibody that interacts with cell surface receptors (Fc receptors) and some proteins of the complement system.
  • This interaction allows antibodies to activate the immune system.

Hinge Regions

  • Found between the CH1 and CH2 regions.
  • Allow antigen-binding “arms” of Ab to flex inward and outward.
  • Rich in cysteines, facilitating heavy-chain dimerization through interchain disulfide bond formation.

Carbohydrate Chains

  • Antibodies are generally glycosylated.
  • Oligosaccharide side chains help “spread” heavy-chain domains apart.
  • Other antibodies have carbohydrates attached to light-chain domains.

Carboxy-Terminal Domains

  • Membrane-bound Ab has three “extra” regions:
    • An extracellular hydrophilic 26 amino acid “spacer”.
    • A 25 amino acid hydrophobic transmembrane segment.
    • A very short three amino acid cytoplasmic tail.
  • Secreted Ab is formed by alternative RNA splicing mechanisms that remove/replace these regions.

T-Cell Receptor (TCR)

  • T-cell receptors (TCRs) are not immunoglobulins, but they have Ig domains and belong to the Ig superfamily of proteins.
  • They possess variable (V) domains and constant (C) domains, similar to Ab molecules.
  • TCRs are heterodimers, mostly with an α (alpha) and a β (beta) chain.
  • They recognize a short peptide carried in the groove of a major histocompatibility complex (MHC) protein on an antigen-presenting cell (APC).
  • TCRs work as part of a complex that includes CD3.

T-Cell Co-receptors CD4 and CD8

  • The T-cell co-receptors CD4 and CD8 also bind the MHC to aid signal transduction.
  • CD4 is a 55 kDa monomer with four extracellular Ig domains and binds to regions on MHC Class II.
  • CD8 is usually a heterodimer with disulfide-linked α and β chains (each 30-38 kDa) and binds to regions on MHC Class I.

T-Cell Accessory Molecules

  • Several different T-cell accessory molecules assist in T-cell signal transduction.
  • Examples include CD4, CD8, CD2 (LFA-2), CD28, CTLA-4, CD45R, and CD5, each with specific ligands and functions.

TCR Heterodimers and Ig Superfamily

  • The domain structures of αβ and γδ TCR heterodimers are similar to those of the Igs, classifying them as members of the Ig superfamily.

TCR Domains and Hypervariable Regions

  • TCR domains, one variable (V) and one constant (C), are structurally homologous to the V and C domains of Igs.
  • The TCR variable domains have three hypervariable regions, equivalent to the complementarity-determining regions (CDRs) in Ig light and heavy chains.
  • Hypervariable regions are usually called CDRs.

γδ TCRs

  • A small subset of T cells carry a γδ (gamma delta) receptor instead of αβ.
  • These cells recognize antigens differently than αβ T cells.
  • The structure of a γδ receptor reacting with a phospholipid antigen reveals significant differences in overall structure compared to αβ receptors, suggesting possible differences in function.
  • γδ receptors have a deep cleft accommodating the microbial phospholipid for which the receptor is specific and do not require MHC presentation.

MHC and CD1

  • MHC Class I presents peptides of 8-10 amino acids.
  • CD1 presents lipid/glycolipid or lipopeptides.
  • MHC Class II presents peptides > 10 amino acids.

MHC Cleft Geometry

  • Peptides are held in the MHC cleft by non-covalent forces.

γδ T Cells and Antigen Recognition

  • While αβ T cells recognize antigen processed and presented in the context of an MHC molecule, γδ T cells do not require either MHC processing or presentation for antigen recognition.

γδ T Cells and Immunity

  • Recognition of antigens common to classes of microorganisms, as well as the ability of γδ TCRs to bind to nonclassical self-MHC molecules suggest a rapid-response role more characteristic of innate than adaptive immunity.

γδ T Cell Characteristics

  • The number of γδ T cells in circulation is small compared with cells that have αβ receptors.
  • The V gene segments of γδ receptors exhibit limited diversity.
  • The majority of γδ cells lack both CD4 and CD8, and the majority express the same γδ-chain pair.

Comparison of αβ and γδ T Cells

  • αβ T cells:
    • Proportion of CD3+ cells: 90-99%
    • TCR V gene germ-line repertoire: Large
    • CD4+ phenotype: ~60%
    • CD8+ phenotype: ~30%
    • CD4+ CD8+ phenotype: <1%
    • CD4- CD8- phenotype: <1%
    • MHC restriction: CD4+: MHC class II, CD8+: MHC class I
    • Ligands: MHC + peptide antigen
  • γδ T cells:
    • Proportion of CD3+ cells: 1-10%
    • TCR V gene germ-line repertoire: Small
    • CD4+ phenotype: <1%
    • CD8+ phenotype: ~30%
    • CD4+ CD8+ phenotype: <1%
    • CD4- CD8- phenotype: ~60%
    • MHC restriction: No MHC restriction
    • Ligands: Phospholipid, intact protein

Role of γδ T Cells

  • γδ cells can secrete a spectrum of chemokines and cytokines, suggesting a regulatory role in recruiting αβ T cells to the site of invasion by pathogens.
  • The recruited αβ T cells would likely display a broad spectrum of receptors.
    *Note: Cytokines are the general category of messenger molecules, while chemokines are a special type of cytokine that direct the migration of white blood cells to infected or damaged tissues.
  • γδ cells may themselves serve as APCs, thus greatly expanding their functional potential.

T-Cell Receptor Complex: TCR-CD3

  • The TCR associates with CD3, forming the TCR-CD3 membrane complex.
    *Recall the B cell receptor complex
  • The accessory molecule participates in signal transduction after interaction of the T cell with antigen; it does not influence interaction with antigen.
  • CD3 is a complex of five invariant polypeptide chains that associate to form three dimers:
    *gamma and epsilon (γε)
    *delta and epsilon (δε)
    *zeta and zeta (ζζ), or zeta and eta (ζη)
    *The ζ and η chains are encoded by the same gene but differ in their carboxyl-terminal ends due to RNA splicing of the primary transcript.
  • 90% of the CD3 complexes incorporate the (ζζ) homodimer; the remainder have the (ζη) heterodimer.

T-Cell Receptor Complex Composition

  • The T-cell receptor complex consists of four dimers:
    1. αβ or γδ TCR heterodimer
    2. gamma and epsilon (γε)
    3. delta and epsilon (δε)
    4. zeta and zeta (ζζ), or zeta and eta (ζη)

CD3 Chain Characteristics

  • The γ, δ, ε chains each contain an immunoglobulin-like extracellular domain followed by a transmembrane region and a cytoplasmic domain of more than 40 amino acids.
  • The ζ chain has a very short external region of only nine amino acids, a transmembrane region, and a long cytoplasmic tail containing 113 amino acids.

Transmembrane Interactions in TCR-CD3 Complex

  • The transmembrane regions of all the CD3 polypeptide chains contain a negatively charged amino acid residue that interacts with one or two positively charged amino acids in the transmembrane region of each TCR chain.

TCR-CD3: Recognition vs. Signaling

  • TCR is responsible for recognition of the antigen.
  • CD3 is responsible for signaling.

T-Cell Accessory Membrane Molecules: CD4 and CD8

  • T cells can be subdivided into two populations according to their expression of CD4 or CD8 membrane molecules.
  • CD4+ T cells recognize antigen that is combined with class II MHC molecules.
  • CD8+ T cells recognize antigen that is combined with class I MHC molecules and function largely as cytotoxic cells.

Affinity of TCR for Peptide-MHC Complexes

  • The affinity (Kd) of TCRs for peptide-MHC complexes is low to moderate.
  • The KD value relates to the concentration of antibody (the concentration of Ab that is required to yield a significant amount of interaction with the target protein) and so the lower the KD value (lower concentration) and thus the higher the affinity of the antibody.

T-Cell Interactions and Cell-Adhesion Molecules

  • T-cell interactions do not depend solely on binding by the TCR.
  • Cell-adhesion molecules strengthen the bond between a T cell and APC or target cell.
  • CD2, LFA-1, CD28, and CD45R bind independently to other ligands on APCs or target cells (these are accessory membrane molecules).