B‐Cell Antigen Receptor: Assembly and Diversification

Abstract

Immunoglobulin (Ig) genes of mammalian B cells undergo two processes that diversify their genomic sequence and structure. The first one concerns the BCR assembly that is achieved through a combinatorial rearrangement of a large number of V, D and J gene segments. This process is essential for B‐cell development and occurs during early B‐cell differentiation in the bone marrow without antigenic challenge. The second process includes somatic hypermutation (SHM), and class switch recombination (CSR), start when the B cell encounters an antigen at the periphery. While the two processes share the action of ubiquitous DNA repair factors to carry out all the steps necessary for creating recombination and mutations in genomic DNA, they are initiated by different lymphoid‐specific factors. On one hand, the action of recombination activating genes 1 and 2 (RAG1 and RAG2) is required for the V(D) J rearrangements and, on the other hand, the action of activation‐induced cytidine deaminase (AID) for both SHM and CSR. The ultimate goal of these diversification mechanisms is to create a dynamic and robust immune response.

Key Concepts

  • V(D)J recombination is a programmed that generates B‐ and T‐cell receptors in vertebrates (BCR and TCR).
  • V(D)J recombination requires precise cutting of the DNA at recombination signal sequences (RSS) followed by rejoining of the resulting termini. Imprecisions during the ends‐joining reaction contribute significantly to increasing the variability of the resulting functional BCR and TCR.
  • RAG1, RAG2 and TdT are the three lymphoid‐specific factors needed for V(D)J recombination and for the junctional diversity observed during lymphoid development.
  • B‐cells provide the antibody‐mediated immune response (humoral immunity). It can generate antibodies to an immense variety of pathogenic antigens.
  • Somatic hypermutation (SHM) and class switch recombination (CSR) occur only in germinal centre B cells.
  • During SHM, DNA repair mechanisms are diverted from their canonical role in preserving genomic integrity to permit high rate of mutations.
  • SHM and CSR both occur in germinal centre in secondary lymphoid tissues and require activation‐induced cytidine deaminase (AID).
  • AID deaminates deoxy‐cytosine on single‐stranded DNA.
  • Ten–eleven translocation (TET) proteins (mainly TET2et TET3) are key regulators of immunoglobulin light chain rearrangement, class switch recombination and plasma cell differentiation.

Keywords: B‐lymphocytes; immunoglobulins; DNA recombination; gene diversification; somatic hypermutation; class switch recombination; DNA repair

Figure 1. Organisation of immunoglobulin genes and mechanisms of diversification. (a) Germline organisation of immunoglobulin genes in mouse. (b) V(D)J recombination during B‐cell development. The first mechanism of diversification occurs during early development of B cells in the bone marrow in an Ag‐independent manner. Heavy chain recombination occurs first, during the early pro‐B cell stage of differentiation. At the large pre‐B cell stage, cells express a pre‐B receptor made of the heavy chain and a surrogate light chain. Later, the light chain starts recombination at the small pre‐B stage. Upon antigen encounter, activated B cells undergo two new mechanisms of diversification: somatic hypermutation (described in Figure ) and class switch (described in Figure ) that take place in peripheral lymphoid tissues.
Figure 2. Overview of D J Recombination mechanism. RAG enzymes bind recombination sequence signal (RSS) following the 12/23 rule. The RAG1/RAG2 heterotetramer along with HMG1 (High Mobility Group 1) bind to each RSS. Rag activity generates double‐strand nicks at the two RSS. The cleaved extremities are maintained nearby and processed by enzymes in the non‐homologous end‐joining pathway. During the DSBs repair mechanism, short deletions or insertions of palindrome base sequences called ‘nucleotides P’ are introduced to increase diversity. The Terminal Deoxynucleotidyl transferase (TdT) adds nucleotides at 3′ end called ‘N nucleotides’, thus increasing further the diversity at IgH locus.
Figure 3. Molecular mechanisms of somatic hypermutation and class switch recombination. (a) Somatic hypermutation induces mutations through two different pathways occurring during different stages of the cell cycle. During G1 phase uracils are recognised by the MSH2/MSH6 complex, then a nick is created on both transcribed and coding strands by Pms2‐Mlh1 or by uracil glycosylase (UNG SMUG1, TDG) along with APEX2 on the coding strand only. Exo1 creates a 30–50 bp gap that is filled by the translesion DNA polymerase η that induces mutations on A/T base pairs. During S phase uracils are eliminated by UNG, creating abasic sites. The recruitment of different translesion DNA polymerases to bypass the abasic sites generates mutations at G/C bases. (b) Models for the generation and repair of programmed DNA DSBs in B cells during CSR. AID target Heavy chains GC‐rich switch (S) regions converting dCs to dUs. Enzymes of the mismatch repair (left) and base excision repair (right) convert the dUs to DSBs which are necessary for the CSR reaction. The DNA ends are subsequently recombined exclusively by NHEJ (see text and Figure for more information).
Figure 4. Organisation of the mouse IgH locus during class switch recombination to IgA.In mouse naïve B cells that express IgM (and IgD) the enhancer Eμ (grey oval) is topologically close to 3′ regulatory region 3′RR (dark grey rectangle). The spatial proximity of those distant sequences is mediated by binding of CTCF (green dots) and loading of cohesin complex (blue ellipse). The diagram only indicates the approximate location of the sites of interaction. During CSR in the germline centre, AID deaminates dC residues in both DNA strands of transcriptionally active S region (Sμ and Sα in the scheme shown). This enzymatic reaction creates uracil bases in the DNA. Processing of these modified bases generates DNA double‐strand breaks in both the donor and recipient S regions that are subsequently joined and ligated by an intrachromosomal deletional recombination. In the shown example, Cα is joined to the expressed VDJ region and the DNA between the two breaks eliminated in circular form. Eμ and 3′RR region are the two major enhancer that regulate expression of IgH genes and CSR. aGLT: alpha germline transcript.
Figure 5. DSB generated during CSR are repaired by NHEJ. Ku heterodimer (dark and light greens) recognises and binds to the DNA breaks to initiate DNA repair. DNA‐PK and MRN complex (blue rounds) are loaded to stabilise DNA extremities in the S‐S synapse. MRN complex recruits ATM (orange pentagon) to phosphorylate 53BP1, Nbs1 or H2A. ATM also induces cell cycle arrest in G2/M. MRE11 (part of MRN complex) partially resects DNA extremities to promote Rev7: shieldin complex loading via Shld2 on ssDNA. Phosphorylated 53BP1 (purple rectangle) prevents over resection necessary for HR and thereby promotes NHEJ. Rif1 and 53BP1 link the shieldin complex to modified histone H2A. After activation by DNA‐PK holoenzyme, XRCC4‐LigaseIV join two incompatible ends. Phosphorylated 53BP1 accumulates at sites of DNA DSBs damage in a manner dependent on RNF8/RNF168 histone ubiquitylation thus facilitating class switch recombination. Adapted from Setiaputra and Durocher .
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Further Reading

Bednarski JJ and Sleckman BP (2019) At the intersection of DNA damage and immune responses. Nature Reviews Immunology 19 (4): 231–242. DOI: 10.1038/s41577‐019‐0135‐6.

Higgins BW, McHeyzer‐Williams LJ, McHeyzer‐Williams MG and Roth D (2019) Programming isotype‐specific plasma cell function. Trends in Immunology 40 (4): 345–357. DOI: 19.01.012 Epub 2019 Mar 4.

Roth DB (2014) V(D)J recombination: mechanism, errors, and fidelity. Microbiology Spectrum 2 (6). DOI: 10.1128/microbiolspec.MDNA3‐0041‐2014.

Schatz DG and Swanson PC (2011) V (D) J recombination: mechanisms of initiation. Annual Review of Genetics 45: 167–202.

Zhang Y, Cheng TC, Huang G, et al. (2019) Transposon molecular domestication and the evolution of the RAG recombinase. Nature 569 (7754): 79–84. DOI: 10.1038/s41586‐019‐1093‐7. Epub 2019 Apr 10 30971819.

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Mohammad, Mahwish M, Guillou, Morwenna Le, Ghamlouch, Hussein, and Aoufouchi, Said(Jan 2020) B‐Cell Antigen Receptor: Assembly and Diversification. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0028602]