Antigen Recognition in the Adaptive Immune System

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Chapter 4

• Antigen Recognition in the Adaptive Immune System

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Ag Recognition

• Specific Ag recognition is the task of 2
  structurally similar types of cell surface
  protein of lymphocytes
• membrane bound Ab on B-cell
• T-cell receptors (TCR) on T-cells
• Function to 1) detect external stimuli and 2)
  trigger response of the cell on which the
  receptor is expressed
• Ag receptors are clonally distributed each
  clone of lymphocytes having a particular
  specificity has a unique receptor different
  from receptors of all other clones

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Ag ReceptorFunctions in Adaptive Immunity
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Function 1

• Ag receptors of B and T cells recognize
  chemically different structures
• B-cells are able to recognize shapes or
  conformations of native proteins
• most T-cells recognize peptide Ag and only on APC
  bound to MHC

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Function 2

• Ag-receptor has domains that are involved in Ag
  recognition
• vary between clones of lymphocytes
• other regions are required for structural
  integrity
• others for effector functions
• All are conserved among all clones
• Ag recognizing portion of the receptors are
  called variable region and conserved portion are
  called constant region
• can maximize variability without changing basic
  structure

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Function 3

• Ag receptors are non-covalently linked to other
  invariant molecules which deliver message to
  inside of cell activation signal
• 2 functions of B-cell and T-cell receptors
  specific Ag recognition and signal transduction
• different polypeptides to do each function
• Receptors with Ag bound will aggregate, causing
  signaling molecules to be close together, enzyme
  will attach on cytoplasmic portion the protein
  and phosphorylation of other proteins complex
  signaling cascade started mediate the lymph
  response

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Function 4

• Ab can be membrane bound or secreted (BCR) but
  TCR are only membrane bound
• Ab or immunoglobulins in the blood neutralize
  microbes and toxins
• recognize microbial Ag and toxins by variable
  domains constant domain binds other molecules
  (receptors on macrophages and complement
  proteins) that participate in eliminating Ag
• BCR functions to initiate humoral immune response
  and secreted Ab eliminates Ag in the effector
  phase of humoral response
• TCR function only in ag recognition and T-cell
  activation do not mediate effector functions

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Antibodies

• 4 polypeptide chains 2 identical heavy (H)
  chains and 2 identical light (L) chains
• L chain is attached to the H chain and the 2 H
  chains are linked by disulfide bonds intrachain
  and interchain

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Antibodies Secreted BCR

• 2 distinct regions
• constant region 1 type of 4 or 5 kinds,
  determines how pathogen is disposed of after
  binding of Ab part L and H chain
• variable region infinite number of forms, both
  arms are identical part L and H chain
• Characteristic folding pattern in each region
  Ig domain and it is found in other proteins in
  and out of the immune system

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Ab Regions

• Variable region is made up of VH and VL and has 3
  hypervariable regions called CDR
• CDR3 is most variable and contributes the most to
  Ag binding
• Fab region is made up of VH,VL, CH1 and CL1
  fragment Ag binding
• Fc is made up of CH2 and CH3 fragment
  crystalline region
• Between Fc and Fab is the hinge region that
  allows the Ab some flexibility for binding Ag
• Fc region is what dictates whether the BCR is
  anchored in the membrane or secreted into the
  intercellular spce

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Proteins of Ab

• Light chain has 2 classes
• ? and ? have different C regions but the same
  function
• Heavy chain has 5 classes
• ?, ?, ?, ? and ? that correspond to IgE, IgA,
  IgM, IgD and IgG respectively
• Many combinations of light and heavy chains
  contribute to the repertoire

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Isotypes

• Ab with different heavy chains that differ in
  properties and effector functions
• All BCR (attached to membrane) are either IgM or
  IgD heavy chains
• it is only after Ag recognition that you will
  have a switch to a different heavy chain and lead
  to secretion from the B-cell heavy chain class
  (isotype) switching
• B cells retain specificity because it is only the
  constant region that is changing, not the
  variable region

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Ag Binding

• Can bind a variety of Ag large macromolecules
  or small chemicals and many shapes
• Ag-Ab reaction is created by H-bonds and charge
  interactions

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Ag Recognition Types

• Recognized components of Ag are called epitopes
  or determinants
• linear determinants AA are all in a row
• conformational determinants dependent on shape
  of Ag and may be AA at distant sites on the
  polypeptide

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Ag-Ab Attraction

• Affinity strength of the attraction between
  epitope and variable region of Ab
• Dissociation constant is the molar concentration
  required to occupy ½ of available Ab the lower
  the KD the higher the affinity
• 1 Ab response is in the range of 10-6 to 10-9
  but the 2 response can boost it to 10-8 to 10-11
  affinity maturation
• Avidity total strength of binding for all
  binding sites and the Ag
• Cross-reaction Ab that recognizes an Ag that is
  structurally similar to Ag it was made against

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BCR Ag Recognition

• BCR will recognize Ag but they require Ig? and
  Ig? as part of the complex
• Ig? and Ig? transmit signals to interior so
  B-cell can be activated
• Monoclonal Ab is one that is made by 1 B-cell to
  recognize 1 Ag
• make by fusing a B-cell with a myeloma cell to
  make a hybridoma that will make just that Ab the
  B-cell is specific for

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T-Cell Receptor for Ag

• ? and ? chain with 1 variable and 1 constant
  region each homologous to Ig V and C regions
• V region has 3 hypervariable or complementary
  regions CDR3 is most important and most
  variable
• Always membrane bound, never secreted
• No class switching or affinity maturation

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TCR Recognition

• ? and ? chain participate in specific
  interactions with MHC and bound peptides
• Recognize as few as 1-3 residues in the
  MHC-associated peptide

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Other T-cell Receptors

• 10 of T-cells have TCR made of ? and ? chains
• have different specificities
• abundant in epithelia
• may recognize a variety of protein/nonprotein Ag
  not displayed by classical MHC molecules
• lt5 are natural killer T-cells
• ??TCR can recognize glycolipid or other
  nonpeptide Ag displayed on non-polymorphic
  MHC-like molecules by a not well understood
  mechanism

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Development of Immune Repertoires

• We get the enormous diversity in immune responses
  because of the maturation of the lymphocyte
• 3 processes involved
• proliferation of immature lymphocyte
• expression of Ag receptor genes
• selection of lymphocytes expressing useful
  receptors
• Common to B-cells (in BM) and to T-cells (in
  thymus)

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Immature Lymphocyte Proliferation

• Maximizes the numbers of cells available to
  express useful Ag receptors and mature into
  functionally mature lymphocytes
• IL-7 stimulated by stromal cells in the BM and
  thymus help lymphocytes proliferate without
  expressing Ag receptors
• later Ag receptors are expresses and then
  regulate signals for proliferation

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Ag Receptor Genes

• Ag receptor gene segments are separated in the
  germline and recombine in lymphocyte maturation
• Various nucleotide sequence changes at site of
  recombination
• Central to lymphocytes maturation

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Maturing Lymphocytes

• Selection at several steps to preserve usefulness
• based on expression of intact Ag receptor
  components and what they recognize, others will
  die by apoptosis
• Positive selection immature T-cells recognize
  Self MHC molecules and then must recognize the
  same MHC to be activated delivers the signals
  for survival and proliferation
• Negative selection eliminates lymphocytes that
  recognize self antigens

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Antigen Receptor Gene Loci

• Initiated by somatic recombination of gene
  segments code for variable region
• Genes are in germline Ig H and L chain and TCR
  ? and ? chain loci each contain multiple V region
  genes (? few 100) and 1 to a few C regions
• Between V and C gene region are several small
  stretches of Joining (J) and Diversity (D) gene
  segments

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B-Cell Receptor

• Lymphocyte progenitor to become a B-cell it must
  associate with recombination of 1 IgVH gene with
  1 D and 1 J segment that is randomly selected
• Immature B-cell has VDJ in the heavy chain locus,
  1 RNA is spliced onto the ?mRNA C region to make
  the IgM receptor on the surface of the B-cell
• Similar things happen in BCR light chain and in
  the TCR ? and ? chains

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Enzymes Involved

• V(D)J recombinase responsible for somatic
  recombination
• Recombinase-activating gene (RAG) -1 and RAG-2
  are protein components that recognize DNA
  flanking the gene sequences, endonuclease cut
  them out and ligases repair the backbone
• Produced only in immature T and B cells
• BCR only on B-cells and TCR only on T-cells
• mechanism unknown

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Diversity by Recombination

• Recombination of different V, D, and J segments
  lead to Combinatorial diversity
• Differences in the nucleotides introduced at the
  junctions leads to Junctional diversity

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Diversity

• Combinatorial is limited by the number of V, D
  and J gene segments
• Junctional is almost unlimited because of the
  different sequences removed

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Enzymes for Diversity

• Exonucleases may remove sequences at ends of V, D
  or J segment and may produce a new sequence
• Terminal deoxyribonucleotidyl transferase (TdT)
  takes sequences not part of the germline and adds
  to the segments of V(D)J recombination forming
  new regions
• Overhanging nucleotides may be filled in with
  P-nucleotide

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Checks and Balances

• Combinatorial and junctional recombination is why
  we have so much diversity in receptors leads to
  CDR3 being able to be used for so many Ag
• May get genes that recombine that dont code a
  functional protein reasons for so many checks
  and balances in lymphocyte maturation

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Maturation and Selection of B-Cell

• Process is random and occurs mainly in the bone
  marrow
• Progenitors commit to be a B-cells proliferate
  under IL-7
• pro-B cells to pre-B cell (get ? heavy chain gene
  recombination, mainly in cytoplasm) happens on
  1 chromosome
• some ? gets to surface with an invariant protein
  similar to light chains to make preBCR complex
• Check point 1 preBCR signals proliferation and
  survival of cells expand pre-B cellswith
  functional ? chains otherwise apoptosis

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Pre-B cell and ? Protein

• Signal 2 processes
• shut off of Ig heavy chain recombination on 2nd
  chromosome Allelic Exclusion
• signals recombination at light chain locus - ?
  then ? - combines with ? chain to make IgM BCR
• BCR on surface signals survival and proliferation
  express complete receptor 2nd check point

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B-Cell Maturation

• Immature B-cells may mature in BM or in
  peripheral lymphoid tissues
• Complex maturation involves co-expression of IgD
  with IgM caused by splicing of the C region
  between ? or ?
• IgMIgD BCR are mature B-cell and can respond to
  antigens
• Repertoire also shaped by negative selection
  binds Ag in BM with high affinity maturation is
  stopped
• may die by apoptosis stops recognition of self
  Ag
• may undergo receptor editing reactive
  recombinase to make a second light chain

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Maturation and Selection of T-cells

• T-cell progenitors migrate from BM to thymus
  maturation occurs here
• Most immature are pro-T-cells or double negative
  T-cells because no CD4 or CD8 expand under IL-7
  influence
• some double negative T-cells undergo TCR ? gene
  recombination by recombinase expressed on
  surface with invariant protein pre-T? to form
  preTCR on pre-T cells, otherwise cell dies
• PreTCR complex sends signals to promote survival,
  proliferation, allelic exclusion of TCR ? locus
  and TCR ? recomination
• no functional ? subunit cell death, cells have
  both CD4 and CD8 double t-cell

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Positive Selection

• Cells that recognize MHC with self-peptide will
  survive but if it doesnt recognize MHC the cell
  will die Self-MHC restricted
• TCRs that recognize MHC class I will keep CD8
  and loose expression of CD4 and MHCII
  recognition, recognize with low avidity
• CD8 cell on activation become CTLs and CD4
  cell become helper cell

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Negative Selection

• Double positive T-cells that recognize
  MHC-peptide complex with strong recognition fie
  by apoptosis high avidity binding

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