Class Switch Recombination: Mechanisms and Key Insights

The immune system relies on antibodies to recognize and neutralize pathogens, but not all antibodies function the same way. B cells undergo class switch recombination (CSR) to modify the constant region of their immunoglobulin genes, enabling them to produce different antibody isotypes without altering antigen specificity. This allows for more effective immune responses tailored to specific threats.

Understanding CSR is essential as it plays a crucial role in adaptive immunity, influencing both defense against infections and susceptibility to autoimmune diseases.

Gene Configuration Of Immunoglobulins

Immunoglobulin (Ig) genes are structured to facilitate antibody diversity, a key feature of adaptive immunity. These genes are located at three primary loci: the immunoglobulin heavy chain (IGH) locus on chromosome 14, the immunoglobulin kappa (IGK) locus on chromosome 2, and the immunoglobulin lambda (IGL) locus on chromosome 22 in humans. CSR occurs within the IGH locus, which encodes the constant (C) regions that determine antibody isotype. Multiple C region genes—Cμ, Cγ, Cα, and Cε—are arranged sequentially, corresponding to different antibody classes (IgM, IgG, IgA, and IgE). This organization enables B cells to switch from producing one isotype to another while maintaining antigen specificity.

The variable (V), diversity (D), and joining (J) gene segments within the IGH locus undergo recombination during early B cell development to generate a functional variable region, a process distinct from CSR. Initially, B cells express IgM and IgD, as these isotypes are encoded by the first constant region genes. Switching to other isotypes occurs through CSR, which involves recombination between switch (S) regions located upstream of each C region gene. These S regions, composed of repetitive DNA sequences, facilitate the deletion of intervening DNA, repositioning a downstream C region gene adjacent to the VDJ segment.

The IGH locus structure is highly conserved across mammals, though variations exist in the number and arrangement of C region genes. Humans have four IgG subclasses (IgG1, IgG2, IgG3, and IgG4), while mice have three. This genetic diversity influences species-specific immune responses, as different IgG subclasses interact differently with Fc receptors and complement proteins. Regulatory elements, such as the 3′ regulatory region (3’RR), enhance transcription of specific C region genes, influencing isotype selection.

DNA Events Leading To Switch Recombination

CSR enables B cells to replace the initially expressed C region of the immunoglobulin heavy chain with a downstream C region, altering the antibody isotype. This process occurs within the IGH locus and involves targeted DNA modifications that facilitate recombination of switch (S) regions. These S regions, which precede each heavy chain C gene (except Cδ), contain repetitive sequences rich in guanine and cytosine, making them susceptible to activation-induced cytidine deaminase (AID)-induced mutations.

CSR begins with transcriptional activation of a specific S region, generating noncoding germline transcripts that precede recombination. AID then deaminates cytosine bases within the transcribed S region, converting them into uracil, creating U:G mismatches. These mismatches are processed by base excision repair and mismatch repair pathways. Uracil DNA glycosylase (UNG) excises uracil bases, generating abasic sites that are cleaved by apurinic/apyrimidinic endonuclease 1 (APE1). The mismatch repair system, including MutS homolog 2 (MSH2) and MutL homolog proteins, introduces additional strand breaks, leading to DNA double-strand breaks (DSBs) at two distinct S regions—typically one upstream of Cμ and another at the target C region.

DSBs are resolved through the non-homologous end joining (NHEJ) repair pathway. The Ku70/Ku80 heterodimer and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) stabilize the breaks and recruit repair factors. Artemis, a DNA endonuclease, trims the DNA ends, while XRCC4 and DNA ligase IV complete the repair by joining the distal S region breaks. This recombination excises the intervening DNA as a circular episome, repositioning the selected C region adjacent to the VDJ exon, enabling expression of a different antibody isotype.

Key Enzymes And Proteins

CSR is orchestrated by enzymes and regulatory proteins that introduce, process, and repair DNA lesions within the IGH locus. These components ensure precise recombination of S regions, allowing B cells to modify antibody isotypes without altering antigen specificity.

Activation-Induced Cytidine Deaminase

AID initiates CSR by deaminating cytosine bases within transcribed S regions, converting them into uracil and creating U:G mismatches that lead to DSBs. AID expression is tightly regulated and occurs predominantly in germinal center B cells after antigen stimulation. Phosphorylation at serine-38 enhances its interaction with replication protein A (RPA), facilitating recruitment to S regions. Studies, such as those published in Nature Immunology (Muramatsu et al., 2000), show that AID-deficient mice fail to undergo CSR, underscoring its essential role. However, AID’s mutagenic potential poses risks, as off-target activity can contribute to genomic instability and oncogenic mutations, particularly in proto-oncogenes like MYC.

DNA Repair Enzymes

Once AID-induced lesions form, DNA repair enzymes process and resolve these breaks. The base excision repair (BER) pathway, involving UNG and APE1, removes uracil residues and generates abasic sites, which are cleaved to create single-strand nicks. The mismatch repair (MMR) system, including MSH2 and MutL homolog 1 (MLH1), amplifies DNA damage by introducing additional strand breaks, facilitating DSB formation. These breaks are repaired through NHEJ, which relies on Ku70/Ku80, DNA-PKcs, and DNA ligase IV to ligate the recombined S regions. Deficiencies in these enzymes, as seen in hyper-IgM syndrome, impair CSR and increase susceptibility to immunodeficiencies.

Transcription Factors

Transcription factors regulate CSR by modulating S region accessibility and transcriptional activity. Nuclear factor kappa B (NF-κB), activated by CD40 signaling, promotes germline transcript production necessary for CSR. Signal transducer and activator of transcription 6 (STAT6) enhances IL-4-mediated switching to IgE and IgG1. B cell-specific activator protein (BSAP), encoded by PAX5, maintains B cell identity and regulates AID expression. Dysregulation of these factors can lead to aberrant CSR, contributing to immune disorders and lymphoproliferative diseases.

Regulatory Switch Regions

Switch (S) regions are highly repetitive DNA sequences upstream of each immunoglobulin heavy chain C region gene, serving as CSR sites. These regions, which span several kilobases, contain tandem repeats rich in guanine and cytosine, making them prone to forming R-loops. In R-loops, the non-template DNA strand remains unpaired while the template strand hybridizes with nascent RNA, enhancing AID accessibility.

S region structure varies between isotypes; for instance, the Sμ region is longer and more complex than Sγ or Sα, influencing recombination efficiency. Epigenetic modifications, including histone acetylation and DNA methylation, regulate their transcriptional activity. Active S regions exhibit histone acetylation at H3K9 and H3K27, while CpG hypermethylation is associated with reduced CSR efficiency. Chromatin remodelers like BRG1 further enhance accessibility by altering nucleosome positioning.

Implications In Autoimmune Conditions

Dysregulated CSR contributes to autoimmune diseases by promoting inappropriate isotype switching and pathogenic antibody production. In systemic lupus erythematosus (SLE), excessive switching to IgG leads to immune complex formation, triggering chronic inflammation and tissue damage. Studies indicate that heightened AID expression correlates with increased somatic hypermutation and CSR activity in SLE patients.

Aberrant CSR is also linked to rheumatoid arthritis (RA) and Sjögren’s syndrome, where altered isotype distributions drive inflammation. RA patients exhibit elevated IgG and IgA rheumatoid factors, contributing to joint degradation, while Sjögren’s syndrome patients show increased IgA and IgG switching in salivary glands, leading to glandular dysfunction. Targeting CSR with therapies like B cell depletion (e.g., rituximab) or modulating AID activity presents potential strategies for mitigating autoimmune responses.

Involvement In Infectious Disease Defense

CSR enhances immune defense by enabling the production of specialized antibody isotypes. IgM, the default antibody, is effective at initial pathogen neutralization but limited in tissue penetration. CSR generates IgG, IgA, and IgE, which provide long-term immunity and specialized defense mechanisms.

IgG mediates systemic infections through opsonization, complement activation, and antibody-dependent cellular cytotoxicity. IgA dominates mucosal immunity, neutralizing pathogens in secretions. IgE is crucial for defense against parasitic infections, triggering mast cell and eosinophil responses. However, excessive IgE responses contribute to allergic conditions, highlighting the need for balanced CSR regulation.