Genetic Basis of Antibody Diversity

GENETIC BASIS OF ANTIBODY DIVERSITY

It has been estimated that an individual is capable of producing up to 10⁹ different antibody molecules. How this vast diversity is generated from a limited number of germline elements has long been one of the most intriguing problems in immunology. There are two possible mechanisms for this variability: either the information is transmitted from gener-ation to generation in the germline, or it is generated somatically during B-lymphocyte differentiation. The following genetic mechanisms have been shown to contribute to the gen-eration of antibody diversity:

1. The existence of a large number of V genes and of a smaller set of D and J segments in the germline DNA, which has probably been generated during evolution as a consequence of environmental pressure. The human VH locus comprises approximately 100 V, 30 D, and 6 J segments.

2. Combinatorial association: as aforementioned, there are at least 100 V-region genes for the heavy chain, and this is probably a conservative estimate. The total number of possible V genes is increased by the fact that any V segment can combine, in principle, with any J and D segments. Imprecise joining of various V-gene segments, creating sequence variation at the points of recombination, augments diversity significantly. In the case of the light chain, the number of V-region genes is estimated as 300, and they can also recombine with different J-region genes. Last, random association of L and H chains plays an important role in increasing diversity. For example, random association of 1000 H chains and 1000 L chains would produce 10⁶ unique antibodies.

3. Somatic mutations were proposed to be a source of antibody diversity in the 1950s. Experimental support for this hypothesis, however, was only obtained three decades later. Comparison of nucleotide sequences from murine embryonic DNA and DNA obtained from plasmocytomas revealed several base changes, suggesting occurrence of mutations during lymphocyte differentiation. There appear to be some special mutational mechanisms involved in immunoglobulin genes since the mutation sites are clustered around the V genes and not around the C genes. In addition to these point mutations, certain enzymes can randomly insert and/or delete DNA bases. Such changes can shift the reading frame for translation (frameshift mutations) so that all codons dis-tal to the mutation are read out of phase and may result in different amino acids, thus adding to the antibody diversity. A large-scale sequencing of H and L chain genes found a much higher proportion of somatically introduced inser-tions and deletions than previously recognized. These insertions and deletions were clustered around the antigen-binding site, thus constituting a major mech-anism of antibody diversity.

Somatic mutations (sometimes termed hypermutations) play a very important role in affinity maturation—production of antibodies with better antigen-binding ability. During the initial exposure to an antigen, rearranged antibodies with appropriate specificity bind to the antigen. Late in the response, random somatic mutations in the rearranged V genes re-sult in the production of antibodies of varying affinities. By a process analogous to natural selection, B cells expressing higher-affinity antibodies are selected to proliferate and those with the lower-affinity antibodies are eliminated.

Additionally, gene conversion, a nonreciprocal exchange of genetic information be-tween genes, has also been shown to contribute to the antibody diversity. It is interesting to note that one of the two recombination-activating genes described earlier, RAG-2, in addi-tion to its synergistic role with RAG-1 in activating VDJ recombination, appears to be in-volved in gene conversion events.

After years of controversy over which mechanism, germline or somatic, is responsi-ble for antibody diversity, it is clear that the involvement of both is necessary.