Somatic Hypermutation and Isotype Switching

Somatic Hypermutation

• Subsequent antigen exposure leads to the accumulation of small point mutations in DNA during rapid cell proliferation after re-stimulation.
• Somatic hypermutation fine-tunes the antibody response and improves the cellular response.
• During cell division, point mutations accumulate, resulting in tighter binding of antigen and receptor.
• Most mutations are nucleotide substitutions rather than deletions or insertions.
• Mutations accumulate in the genes contributing to the epitope binding regions.
• These mutations can change the affinity of the receptor, therefore improving affinity maturation and cellular immune responsiveness.

Isotype Switching

• Isotype switching allows the adaptive immune system to produce antibodies with identical specificity that can initiate different immune responses.
• The hyper-variable region is the warhead, and the constant region is the rocket.
• B cells can manufacture a single type of warhead but can place it on different rockets (constant regions).
• A switch is located upstream from the constant region on the heavy chain.
• As the DNA loop is removed, rejoining occurs at a new region, recombining a unique constant region to the VDJ segment.
• Isotype switching generates warheads positioned on different rockets to specialize the immune attack.
• The rocket determines whether the antibody activates complement, is secreted in the lumen or on the mucous membrane, or enters a specific tissue.
• Isotype switching may involve deletion of DNA to recombine VDJ segments or serial reactivation to memory B cells and shorter deletions.

The Process of Isotype Switching

• Initial Expression: A naïve B cell first expresses IgM and IgD. using the μ (mu) and δ (delta) constant region genes, respectively.
• The variable region is the same, but the constant region initially produces IgM or IgD.
• Activation: A B cell receives signals that induce isotype switching when activated by an antigen and T cell help.
• Cytokines from helper T cells direct the B cell to switch to a specific antibody class (e.g., IgG, IgA, or IgE).
• DNA Recombination: The switch region upstream of each constant region gene contains sequences recognized by activation-induced cytidine deaminase (AID).
• AID causes DNA double-strand breaks in the switch regions.
• The DNA loop between the initial constant region gene and the target constant region gene is excised.
• Rejoining: The switch regions corresponding to the new constant region align and are rejoined. The heavy chain gene is now recombined with the new constant region.
• The B cell will start producing the new isotype of the antibody while retaining the same variable region.
• Outcome: The B cell now produces antibodies of the new isotype with the same antigen specificity but with different functional properties.

Somatic Hypermutation and Class Switch Recombination

• Somatic hypermutation involves rapid, single base pair changes introduced into the variable region of the antibody, affecting both the heavy and light chains.
• The consequence of hyper mutations is a change in the affinity of the antibody.
• Class switch recombination switches the antibody class, which is defined by the heavy chain residue, only affecting the heavy chain.
• A virgin B cell makes the antibody class that is first in line -- IgM and IgD.
• Class switch recombination is driven by a recombinase enzyme complex, that makes two cuts and then recombines the DNA.
• The first cut is fixed right before the mu and delta gene, while the second cut depends on the cytokines in the germinal center.
• A lot of interferon gamma around results in the IgG isotype.
• High levels of TGF beta leads to the IgA isotype.
• Cytokines produced by the T helper cell determine which isotype is produced.

Comparison of Somatic Hypermutation and Ig Isotype Switching

• Somatic Hypermutation Location: Occurs in the variable regions (VH and VL) of the immunoglobulin genes.
  • Activation-Induced Cytidine Deaminase (AID): Initiates SHM by introducing point mutations (cytosine to uracil deamination) in the variable regions.
  • Point Mutations: Deamination leads to uracil formation, resulting in errors during repair, causing point mutations.
  • Affinity Maturation: B cells with higher affinity antibodies have a selective advantage, surviving and contributing to the immune response.
  • Primary Function: To increase the affinity of antibodies for their target antigens in germinal centers.
• Isotype Switching Location: Occurs in the constant region (C region) of the immunoglobulin heavy chain gene.
  • Activation-Induced Cytidine Deaminase (AID): Crucial for initiating isotype switching.
  • DNA Recombination: AID induces double-strand breaks in the DNA at switch regions within the constant region.
  • Switching to Different Isotypes: Recombination replaces one constant region (e.g., Cμ for IgM) with another (e.g., Cγ for IgG).
  • Role: Allows B cells to change the antibody class while maintaining specificity.
  • Function: Adaptation of Effector Functions: Enables B cells to produce antibodies with effector functions appropriate for the immune response needed.

Allelic Exclusion

• B cells are deployed to contain both maternal and paternal chromosomes.
• Allelic exclusion ensures B cells only possess a single antigenic specificity.
• A single B cell/plasma cell can express only kappa variable in constant alleles, or the lambda variable in constant light chain alleles.
• Alleles originate to the exclusion of all others, from maternal or paternal origin
• Allelic exclusion ensures the antigenic specificity of the B cell.
• Each B cell/plasma cell has four light chain gene clusters.two on kappa chromosome 2, two on lambda chromosome 22.
• The heavy chain is located on chromosome 14.
• Only one light chain, from either maternal or paternal sources, is contributed to the immunoglobulin.
• Likewise, the heavy chain will originate from either the mother or the father, not both.

Junctional Diversity

• Diversity is a major theme in immunology.
• Additional receptor variation comes from junctional diversity.
• Exposed ends of gene segments undergo rearrangements through the addition or deletion of nucleotides by TDT.
• Junctional diversity leads to productive combinations that encode alternative amino acids, increasing antibody and receptor diversity.
• TDT can add or remove nucleotides randomly.
• Coding joints fall within the third hyper-variable region in IG heavy and light chains, important for recognition diversity.

Junctional diversity.

• The variable region of the heavy chain of an antibody consists of V, D, and J gene complexes.
• The constant region consists of genes at the C complex.
• During B cell maturation, the DNA that codes for antibodies is rearranged.
• The DNA between a D gene complex and a J gene complex is removed, enabling the D gene and the J gene to combine.
• The DNA between the neighboring V gene complex and the new DJ gene is removed, and the V gene and the new DJ gene combine.
• The new VDJ gene, which codes for the variable region, and a C gene, which codes for the constant region, are combined (somatic recombination).
• The cutting and slicing of the genes is conducted by an enzyme, during which existing nucleotides may be lost and new, non-encoded nucleotides may be added at the gene junction (junctional diversity )
• Because rearrangements of genes cannot be reversed, an individual B cell can only produce one type of antibody.

Comparison of Allelic Exclusion and Junctional Diversity

• Allelic Exclusion: Ensures each B cell expresses only one functional allele of the immunoglobulin heavy and light chain genes.

  • Immunoglobulin Gene Rearrangement: V(ariable), D(iversity), and J(oining) immunoglobulin genes rearrange to create different antibody specificities.
  • Pre-BCR Formation: After heavy gene rearrangement, pre B-Cell receptors express on the the B-cell surface. The rearranged heavy chain and surrogate chain make up the BCR but not a rearranged light chain.
  • Positive Selection: B cells that express Pre-BCR are selected for development.
  • Light Chain Gene Rearrangement: light Chain genes have been rearranged once allelic expression of the Heavy Chain have occurred, they complete the final B-Cell receptor.
  • Role: Ensures monoallelic expression of immunoglobulin Heavy and light chain genes in a single B-Cell.
  • Function: Specificity: The antibody has unique antigenic specificities due to the single allele expression of the Heavy and Light chains genes of the B-Cells. This allows for recognition of broad pathogens.

• Junctional Diversity: Adds/Removes nucleotide to or from V (variable), D (diversity), and J (joining) to the Heavy and Light Chains Immunoglobulins.

  • Random Addition or Removal of Nucleotides: Terminal deoxynucleotidyl transferase (TdT) helps with inserting or deleting nucleotides at the joints of the chains.
  • Combinatorial Variety: The variability of the variable region of the chains increases significantly.
  • Variable Region Formulation: The variable regions are distinct due to the variability introduce by the junctional diversity. This affect the antibody bonding specificities.
  • Role: Junctional Diversity further alter Variable regions of the Heavy and Light chains during gene rearrangement process.
  • Function: Enhanced diversity is achieved with combination of Junctional diversity, and combinatorial diversity , this allows multiple antibodies to be created by B-Cells to deal with multiple different immunogen bond specificities. Enabling the immune system to react and adapt to different antigens.