BIOCHEMISTRY - L. Stryer - 1984

VOLUME 3

PART V. MOLECULAR PHYSIOLOGY

CHAPTER 33. IMMUNOGLOBULINS

33.22. Joining of V and J genes in different reading frames also contributes to antibody diversity

A second surprising discovery was that a set of five J genes directs the synthesis of not five, but a much larger number of Amino acid sequences for the corresponding light-chain regions. Amino Acid and nucleotide sequence analyses revealed that the recombination of V and J genes is not completely precise. As it turns out, the recombination of these genes can occur at various base positions near the codon specifying residue 95 (Fig. 33.26). Consequently, variations in the reading frames during V-J joining provide an additional contribution to antibody diversity in the Organism. It seems that The Immune System delights in minor errors!

Class="center">Fig. 33.26. Imprecision in the joining site of V and J genes serves as an additional source of antibody diversity

In any given lymphocyte, only one of the two allelic genes is expressed. Consequently, all antigen-binding sites produced by an individual Cell are identical. The structural basis for this selective Gene Expression (known as allelic exclusion) has been uncovered. As shown by restriction fragment analysis, the incomplete V gene is correctly joined to the J gene on only one of the two homologous Chromosomes; only this properly recombined immunoglobulin gene is expressed.

33.23. mRNAs for L and H chains are formed by splicing primary transcription products

Light κ-chain mRNA contains about 1250 bases (Fig. 33.27). Like other eukaryotic mRNAs, it contains a poly(A) tail attached to the 3' end and untranslated sequences at the 3' and 5' ends. The leader sequence near the 5' end of the κ-chain mRNA encodes the hydrophobic N-terminal region of the synthesized κ-chain. The resulting signal sequence (Fig. 33.28) directs the ribosome to the Endoplasmic reticulum and determines the ability of the newly synthesized immunoglobulin polypeptide chain to cross the ER membrane into the lumen of its tubules (Sec. 29.30). Subsequently, on the inner surface of the ER membrane, the signal sequence is cleaved by a peptidase. The Variable and constant Regions of the light chain are encoded by an adjacent region of the mRNA that immediately follows the leader sequence.

Fig. 33.27. Structure OF THE mRNA encoding the L chain

Fig. 33.28. Signal sequence of the newly synthesized L chain. Hydrophobic residues are highlighted in yellow

The primary transcript from which this mRNA is formed contains two intervening sequences (Fig. 33.29). One of them separates the leader sequence from the beginning of the mRNA encoding the variable region, and the second is located between the distal end of the sequence complementary to the J gene and THE START OF the sequence complementary to the C gene (i.e., encoding the constant region). During the Processing of the primary transcript into mRNA, these intervening sequences are removed. Curiously, the J gene carries information for Two Types of splicing: one at the DNA level (fusion of V and J genes) and the other at the RNA level (joining of J and C regions).

Fig. 33.29. Primary transcript precursor of L-chain mRNA

What is The structure of the gene encoding the heavy chain? Recall that the H chain consists of four domains: V_H, C_H1, C_H2, and C_H3. The DNA fragment specifying the constant region of the IgG heavy chain has recently been cloned. Electron microscopic studies have shown that C_H1, C_H2, and C_H3 are encoded by separate DNA segments (Fig. 33.30). Yet another DNA segment encodes the hinge region between C_H1 and C_H2. Overall, the domain structure of IMMUNOGLOBULINS (Sec. 33.14) reflects the architecture of their respective genes. Through splicing, organisms have proved capa

ble of generating new Proteins during evolution by combining DNA segments that encode different domains.

Fig. 33.30. Three domains of the heavy-chain constant (C) region and the hinge region are encoded by different gene segments

33.24. Different classes of antibodies are formed as a result of V_H gene switching

As already mentioned, There are five classes of immunoglobulins. Antibody-producing Cells initially synthesize IgM and subsequently switch to IgG, IgA, IgD, or IgE of the same Specificity. During this switch from IgM to another immunoglobulin class, the light chain remains unchanged. Moreover, the variable region of the heavy chain also remains unchanged. Only the constant region of the heavy chain changes, which is why this stage of antibody-producing Cell Differentiation is referred to as C_H switching (Fig. 33.31).

Fig. 33.31. Synthesis of various immunoglobulin classes. As a result of the V_H region gene first joining with the Cμ region gene and subsequently with another C-region gene, heavy chains of different immunoglobulin classes are formed

In mouse embryonic cells, the genes encoding the constant regions of the μ-, γ-, and α-heavy chains (designated as Cμ, Cγ, and Cα, respectively) are arranged in a row, one after another (Fig. 33.32). As it turned out, there are four constant region genes for γ-chains, which is in full agreement with genetic analysis data revealing four subclasses of IgG. Adjacent to the Cμ gene is a set of tandemly arranged J genes encoding the final hypervariable segment of the variable region. A complete IgM heavy chain gene is formed by the translocation of the V_H gene to the J_H gene (Fig. 33.33). As a result of this translocation, the V_H, J_H, and Cμ genes are joined into a functionally single gene. The intervening sequences between the leader segment and the start of the variable region gene, between the end of the J_H gene and the start of the Cμ gene, as well as within the Cμ gene itself, are spliced out during The conversion of the primary transcript into mRNA for the μ-chain.

Fig. 33.32. The constant region genes for the μ-, γ-, and α-chains are arranged in a row, one after another. The positions of the Cδ and Cε genes have not yet been established

Fig. 33.33. Joining of the V_H and J_H genes results in The formation of the gene encoding the μ-chain

As restriction endonuclease Cleavage analysis of DNA from embryonic and myeloma cells has shown, switch recombination occurs at the DNA level rather than the RNA level. For instance, during switching from IgM to IgA, the V_HJ_H gene located next to the Cμ gene is translocated to a region adjacent to the C_a gene (Fig. 33.34). During this recombination, the genes between Cμ and C_a form a loop and are excised. It is possible that the DNA segments undergoing recombination carry a palindromic sequence. It is precisely the translocation of the entire V_HJ_H gene that explains the fact that the IgA produced by a given cell is identical in antigenic specificity to the IgM synthesized by the same cell at an earlier developmental stage. How The Cell selects one of several C_H genes for translocation remains unknown. The Biological Significance of C_H switching lies in the fact that the entire antigen-recognizing domain (the variable domain) is shifted from the initial constant region (Cμ) to other constant regions that encode polypeptide chains with different effector Functions.

Fig. 33.34. Structural basis of C_H switching. As a result of intrachromosomal recombination, the V_HJ_H gene, previously located near the Cμ gene, becomes positioned adjacent to the C_a gene