The immune system recognizes foreign molecules (antigens) through special receptors which enables them to elicit an immune response against them. The adaptive immune system has B-lymphocytes as one of the main players which produce antibodies as their antigen recognizing molecules. Each antibody/immunoglobulin molecule has a unique recognition site which is specific for a particular epitope. Since the immune system is capable of protecting the organism against an infinite number of antigens, a vast receptor repertoire is necessary. Complex molecular genetic mechanisms have evolved to create this enormous diversity in antibody molecules.
Each antibody molecule is composed of two heavy and two light chains each of which has a variable and a constant domain. The genes encoding the heavy and light chains are organized into three clusters - λ (chromosome 22) and κ (chromosome 2) for the light chain and the heavy chain domain (chromosome 14). The variable region which has the antigen binding properties is encoded by the V region for both the heavy and the light chains. The C domain codes for the constant region of both chains and gives the molecule its biological properties. Complete chains are produced by joining the V region to the C region by RNA splicing after transcription.
The V region contains multiple copies of several gene segments in germline DNA which undergo rearrangement and form the complete V gene exon. For the light chain, the V segment codes for the first 95-101 amino acids followed by the J (joining) segment which codes for the next 13 amino acids (approximately). The heavy chain locus has an additional D (diversity) segment between the V and J segment.
B-cells are continuously produced from undifferentiated haematopoietic stem cells in the fetal liver and the bone marrow in adults which provides a specialized microenvironment for its development. The developmental stages are marked by gene rearrangement and expression of specific cell surface markers. Rearrangement occurs in a fixed sequence and acts as an important checkpoint during the B-cell development.
• The earliest cell in B cell lineage development is the early pro-B-cell derived from the stem cell. The DH-JH rearrangement takes place in this stage at the both the alleles which is unique and stands true for only this rearrangement step. Since the failure rates are high in the subsequent steps, a rearrangement in both the alleles is an advantage. In humans, most D-JH rearrangements are successful since the D genes can be read in any of the three reading frames without encountering a stop codon.
• A productive rearrangement signals the cell to proceed to the late pro-B-cell stage where the V-DJH rearrangement takes place. 45% of the pro-B-cells are lost at this stage due to out-of-frame V-DJH rearrangements.
• Successful rearrangement of the VDJH segment leads to a transient expression of the µ heavy chain which is expressed in the cytoplasm and to an extent on the cell surface with a surrogate light chain (λ5 and VpreB) as the pre-B-cell receptor. The pre-B-cell receptor stimulates the cell to become a large pre-B-cell and also halts the heavy chain rearrangement. The large pro-B-cell proliferates actively and greatly expands the population.
• After proliferation the cell becomes a resting small pre-B-cell in which the expression of the surrogate light chain stops and the µ heavy chain remains in the cytoplasm by itself. This stage sees the rearrangement of the light chain genes which is due to the expression of RAG proteins. It is observed that the κ locus rearranges before the λ locus but it is not a compulsion. Chances of a productive rearrangement in the light chain loci are higher because of the scope for repeated rearrangements of the unused V and J segments and the presence of two light chain loci.
• Successful rearrangement of both the chains and their joining leads to the expression of a complete IgM molecule on the cell surface of an immature B-cell.
• A mature B-cell/naive B-cell expresses both IgM and IgD on the cell surface produced after mRNA splicing of the heavy chain gene transcript. These cells circulate through the lymphoid system and get activated when encountered by the appropriate antigen.
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