6.1 Building a receptor #
6.1.1 Segments, signals, and the 12/23 rule #
Antibody genes are inherited as libraries, not as intact genes. The heavy-chain locus on human chromosome 14 has V, D, and J segments upstream of the constant genes. ฮบ on chromosome 2 and ฮป on chromosome 22 have V and J only. Tonegawa showed that developing lymphocytes assemble one combination by DNA rearrangement. RAG1 and RAG2 cut at recombination signals: a heptamer, a 12- or 23-base-pair spacer, and a nonamer. The 12/23 rule joins a 12-spacer only to a 23-spacer, which prevents VโV or JโJ joins and forces heavy-chain order VโDโJ. Coding ends are opened from a hairpin and joined imprecisely; signal ends are joined precisely and usually discarded as a circle. Molecular Biology owns the chemistry of that joining. The immunological result is a new coding joint.
6.1.2 Where diversity actually comes from #
Combinatorial choice of segments is real, and junctional diversity is larger. Opening the hairpin can leave a short palindromic addition, nucleases remove bases, and terminal deoxynucleotidyl transferase adds non-templated N nucleotides. The junction falls in CDR3, so two cells that chose the same segments still differ there. TdT is strong at the heavy chain, weak at the light chain, and scarce in foetal life, so foetal receptors are less random. Many joints are out of frame. The cell tries the other allele or dies.
6.1.3 Order and allelic exclusion #
In the pro-B cell, D joins J and then V joins DJ. A productive ฮผ chain pairs with the surrogate light chain VpreB and ฮป5 as the pre-B-cell receptor. Success proliferates the cell and stops rearrangement of the other heavy-chain allele: allelic exclusion, one heavy-chain specificity per clone. Failure of both alleles kills the cell. Pre-B cells then rearrange ฮบ before ฮป. A productive light chain assembles surface IgM and stops further light-chain rearrangement. The immature B cell has one receptor and has not met foreign antigen.
6.2 Changing the receptor after antigen #
The study of Changing the receptor after antigen provides fundamental principles governing cellular architecture, molecular interactions, and physiological regulation.
Antigen Presentation
6.2.1 Somatic hypermutation and affinity maturation #
Early secreted IgM uses the receptor the marrow released. In a T-dependent response, germinal-centre B cells mutate variable regions. Activation-induced deaminase deaminates cytosine to uracil in single-stranded DNA at the immunoglobulin locus, and error-prone repair spreads mutations to neighbouring bases, at about one mutation per thousand base pairs each division. The T-cell receptor is not mutated this way. In the light zone, the B cell whose receptor captures more antigen from immune complexes on follicular dendritic cells presents more peptide and wins more help from T follicular helpers. Winners can return to the dark zone and mutate again. Affinity of the chosen clones often rises by two or more orders of magnitude.
6.2.2 Class-switch recombination #
ฮผ is the first constant gene downstream of VDJ, so the first secreted antibody is IgM. Class-switch recombination places the same VDJ next to a downstream constant gene and deletes the DNA between. Specificity is unchanged and the Fc changes. The deletion is irreversible, unlike IgM/IgD co-expression, which was splicing. AID deaminates switch regions upstream of each constant gene except ฮด, and the resulting breaks are joined. Cytokines choose the isotype by opening a switch region: in humans, IL-4 favours IgG4 and IgE, and TGF-ฮฒ favours IgA. IL-21 supports switching and, with IL-4, supports IgE. In the mouse, IL-4 instead drives IgG1 and IgE. CD40 ligand plus cytokine turns AID on. Without CD40 ligand, IgM remains and switched isotypes do not appear.
Mhc Class 2
6.3 Checkpoints, help, and the germinal centre #
The study of Checkpoints, help, and the germinal centre provides fundamental principles governing cellular architecture, molecular interactions, and physiological regulation.
Immune Checkpoints Of Immunosuppressive Actions Associated With Breast Cancer
6.3.1 Stages and tolerance #
| Stage | Receptor event | What tolerance does |
|---|---|---|
| Pro-B | Heavy-chain DโJ, then VโDJ | No receptor yet |
| Pre-B | Pre-BCR, then light-chain rearrangement | Allelic exclusion |
| Immature B | Surface IgM in the marrow | Editing, deletion, or anergy if self-reactive |
| Transitional | Leaving for the spleen | A second filter |
| Mature naive | IgM plus IgD in follicles | Needs help; antigen alone is not enough |
| Germinal-centre and plasma cell | Mutation and switching | Competition for antigen and for T-cell help |
An immature B cell that binds multivalent self antigen in the marrow re-expresses RAG and tries a new light chain. That is receptor editing; if it fails, the cell dies. Soluble self antigen more often anergises the cell: it persists with little surface IgM and responds poorly. A second filter acts on transitional cells in the spleen. In the periphery, antigen without T-cell help and without innate pattern signals does not produce a useful response, which is how abundant self proteins that never arrive linked to a foreign peptide are ignored.
6.3.2 T-dependent and T-independent antigens #
Protein antigens are T-dependent. The B cell internalises its epitope and presents peptides; a primed CD4 cell specific for a linked peptide supplies CD40 ligand and cytokines, and the B cell can switch, mutate, and become memory or a long-lived plasma cell. T-independent antigens skip that help. TI-1 antigens also engage an innate receptor and, at high dose, can activate many clones. Lipopolysaccharide via TLR4 is the mouse example; human B cells express little TLR4, so that example does not transfer unchanged. TI-2 antigens are repetitive, classically capsular polysaccharides, and cross-link many receptors. Marginal-zone and B-1 cells respond with largely unmutated IgM and poor memory, and children under about two years respond weakly. That failure is why pure polysaccharide vaccines were rebuilt as conjugates.
6.3.3 Germinal centres #
Some activated B cells become short-lived extrafollicular plasmablasts and supply early, mostly unmutated IgM. Others enter the follicle with T follicular helpers and build a germinal centre. Centroblasts in the dark zone divide and express AID. Centrocytes in the light zone compete for antigen on follicular dendritic cells and for IL-21 and CD40 ligand from T follicular helpers. Winners leave as memory B cells or as long-lived plasma cells that settle in marrow and secrete for years. Plasma cells have lost surface immunoglobulin and are no longer selected by antigen binding.
6.4 Key terms, summary, and self-check #
Key terms. V(D)J; RAG; 12/23 rule; junctional diversity; TdT; allelic exclusion; pre-B-cell receptor; receptor editing; AID; somatic hypermutation; class switch; T-independent antigen; germinal centre; follicular dendritic cell.
Summary. RAG joins V, D, and J under the 12/23 rule, and junctional additions make CDR3 the diverse loop. Heavy chain rearranges before light chain, and success shuts the other allele. Self-reactive immature B cells edit or die. After a protein antigen, AID mutates the variable region and switches the constant gene, and germinal centres keep the higher-affinity mutants. Pure polysaccharides bypass T-cell help and do not make that memory.
Self-check.
1. What does the 12/23 rule prevent, and which enzyme makes the cut?
2. Where do N nucleotides come from, and why does CDR3 diversify more than CDR1?
3. Which chain rearranges first, and what does allelic exclusion achieve?
4. What reaction does AID catalyse, and how can it yield either a point mutation or a new isotype?
5. Why does a pure polysaccharide fail to produce switched, affinity-matured memory? What does a germinal centre add that an extrafollicular focus does not?
