**Generation of Antibody Diversity**

**Generation of Antibody Diversity**

#### Oliver Backhaus Oliver Backhaus Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.72818

#### **Abstract**

Because of the huge diversity, the immunoglobulin repertoire cannot be encoded by static genes, which would explode the genomic capacity comprising about 20,000–25,000 human genes. The immunoglobulin repertoire is provided by the process of somatic germ line recombination, which is the only controlled alteration of the genomic DNA after meiosis. It takes place in mammalian B lymphocyte (B cells) precursors in the bone marrow. The genome germ line sequence of undeveloped B cells is organized in gene segments and compromise V (variable), D (diversity), and J (joining) gene segments constituting the variable domain of the heavy chain and only V and J genes for building up the variable domain of the light chain. The rearrangement of the variable region follows a strict order. The following processes that participate in the generation of antibody diversity were summarized—allelic, combinational, and junctional diversity, pairing of IgH and IgL, and receptor editing—which all together produce the primary antigen repertoire (pre-antigen stimulation). When a B cell encounters a foreign antigen, affinity maturation and class switch are induced. Thereby the antibody repertoire increases. The resulting secondary immunoglobulin repertoire reveals in humans at least 1011 specificities for different antigens.

DOI: 10.5772/intechopen.72818

**Keywords:** antibody diversity, somatic recombination, somatic hypermutation, class-switch recombination, allelic exclusion, B-cell receptor editing, pairing of VH and VL, germinal center

### **1. Introduction**

The immune system is a complex system, comprising different organs and many specialized cell types, which are carrying out their development, maturation, and pathogen recognition at various sides in the body. The immune system has two major approaches to recognize and attack pathogens. The first is the innate immunity followed by the delayed adaptive immune response, based on specific antigen recognition receptors. The innate immune system is

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© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons

nonspecific and uses general pathogen recognition mechanisms, through pathogen-associated molecular patterns (PAMPs) recognized by cell surface or intracellular pattern recognition receptors (PRRs), such as toll-like receptors or NOD-like receptors (NLRs) and RIG-I-like receptors (RLRs) [1]. Cell types of the innate immunity are monocytes/macrophages, dendritic cells, mast cells, natural killer cells, granulocytes, B1 cells, and innate lymphoid cells (ILCs). Although it lacks specificity, it can react immediately on the invading pathogens and activates the adaptive immune system by presentation of the foreign antigen peptides.

**2. The primary antibody repertoire**

**2.1. Combinatorial diversity of immunoglobulins**

comprises 38–46 genes, which varies between individuals.

modified from IMGT [5]).

**Ig chain Chromosomal** 

**location**

**Gene locus**

1

Before immature naive B cells encounter a foreign antigen, their genomic sequence is rearranged by a well-controlled process, called somatic DNA recombination. This process is unique in lymphocytes, and except of the meiosis in the gametes, this is the only DNA recombination of somatic cells [4]. Before B cells leave the bone marrow to the secondary lymphatic organs, somatic DNA recombination takes place. The sum of all B lymphocytes in an individuum, producing different antibodies with different specificities and affinities, is designated as the antibody repertoire. In humans, the antibody repertoire consists of at least 1011 specificities [4]. The number varies and is limited by the total number of B cells and encountered antigens of an individuum. The immunoglobulin loci contain gene fragments to build up all immunoglobulin variable domains of the heavy and light chain. The different immunoglobulin loci are located on different chromosomes (Chr), the heavy chain on Chr14, the kappa light chain on Chr2, and the lambda light chain on Chr22. In contrast to the light chain loci, the heavy chain locus has several constant regions; each represents a different immunoglobulin isotype, e.g., IgM, IgD, IgG1, Ig2a, IgG2b, IgG3, IgE, and IgA in mice. The gene segments consist of different germ line sequences. For example, the variable gene locus of the heavy chain

Besides different germ line segments, there exist a relative large number of pseudogenes of which some can undergo recombination leading to a nonfunctional variable region. An overview of the number of gene segments in the respective gene locus is given in **Table 1** (slightly

The light chain loci have only variable (V) and joining (J) gene segments, whereby the heavy chain locus additionally has a diversity (D) gene segment, which lay between the V and J genes of the heavy chain variable region. One of each gene segment is randomly selected by the RAG1/RAG2 recombinase and joined together to form the variable region (**Figure 2c**) as shown as example with the variable region of the λ light chain. The recombination steps of the V region follow a strict order. The variable light chain recombines first with the V-J segments. Afterward the constant (C) domain is joined through RNA splicing of the primary RNA to the variable region. The construction of the V region of the heavy chain begins with

> **Locus size (kb)**

IGH Heavy chain 14q32.33 1250 38–46 23 6 9 IGK κ Light chain 2p11.2 18201 34–38 0 5 1 IGL λ Light chain 22q11.2 1050 29–33 0 4–5 4–5

**Table 1.** Number of functional human immunoglobulin gene segments in the heavy and light chain locus.

In one known haplotype, the locus size is reduced to 500 kb comprising only 17–19 IGκV genes.

**Variable (V)**

**Immunoglobulin (Ig) gene segments**

**Diversity (D)**

**Joining (J)**

Generation of Antibody Diversity

3

http://dx.doi.org/10.5772/intechopen.72818

**Constant (C)**

The adaptive immune system needs to be activated and primed by the antigen and therefore acts delayed from the initial pathogen attack. It is based mainly on two cell types, the B cell and the T cell. Both cell types express specific receptors on their cell surface for pathogen recognition. Many different B- and T-cell clones exist in parallel inside the body, and each has a different receptor specificity to the antigen. These receptors were called B-cell receptor (BCR) and T-cell receptor (TCR). It is remarkable that despite the relatively small genome size of approximately 20,000–25,000 human genes [2, 3], the human body can produce an antibody repertoire which can recognize almost every possible antigenic structure. Of course, this cannot be achieved by encoding the antigen receptor specificity directly in the genome sequence.

The huge B-cell diversity is generated by a complex multistep process, starting in the bone marrow and ending up in the peripheral lymphoid tissues, such as lymph nodes, spleen, or mucosal lymphoid tissue. In the maturation of functional BCR or TCR, the antigen receptor genes were rearranged from many different possible gene segments to form a full receptor. In each step, the receptor is tested for functionality and excluded when it reveals self-antigen reactivity in order to prevent autoimmunity and making the immune system self-tolerant.

The B-cell maturation occurs inside the bone marrow before the B cells migrate to peripheral lymphoid tissues. On the contrary to B cells, T-cell progenitors migrate to the thymus to differentiate and to mature. After their maturation, B and T cells meet again in lymph nodes. In the germinal centers of the lymph node, antigens were presented to the B cells through antigen-presenting cells, particularly through follicular dendritic cells (FDC). In response to a foreign pathogen, B cells with the highest antigen affinity were selected from a pool of different BCR clones. This process is organized in a form of a repetitive cycle inside of the dark and light zone of a germinal center of the lymph node and is known as the cyclic reentry model (**Figure 4**).

An essential part of the cycle is the BCR affinity maturation of the B cells. It begins with the tight controlled somatic hypermutation (SHM), particularly in the variable regions of the light and heavy chain of the antigen receptor and is only active in the dark zone of the germinal center. This process creates BCRs with higher affinity, whereby the mutations which produced very low or nonfunctional receptors were excluded. Finally, high-affinity B cells differentiate either into plasma cells, which start to produce secreted antibodies with the same specificity as the BCR, or they differentiate into memory cells, conferring lifelong immunity.

This chapter will discuss in detail the different steps and processes, which contribute to the high diversity of B cells. Many steps are similar for the generation of T-cell receptor diversity and were not covered by this article.
