Chapter 5. Generation of lymphocyte antigen receptors

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1 Chapter 5 Generation of lymphocyte antigen receptors

2 Structural variation in Ig constant regions Isotype: different class of Ig Heavy-chain C regions are encoded in separate genes Initially, only two of C and C genes are expressed Thus, naïve B cells have both IgM and IgD on surface During the course of Ab response, activated B cells can switch to expression of individual C H genes by a process of somatic recombination, isotype switching

3 Different classes of Igs are distinguished by structure of their heavy-chain constant regions 5 main isotypes of Ig: IgM, D, G, E, A IgG: IgG1, 2, 3, 4 IgA: IgA1, 2 Sequence differences between Ig H chains cause various isotypes to differ in several characteristic respects - Number and location of S-S bonds - Number of attached oligosaccharide moieties - Number of C domains - Length of hinge region Structural variation in Ig constant regions

4 The properties of the human immunoglobulin isotypes

5 The properties of the human immunoglobulin isotypes

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7 The constant region confers functional specialization on the antibody C region is essential for recruiting other cells and molecules to destroy pathogens to which Ab has bound Fc of Abs have 3 main effector functions Fc of certain isotypes are recognized by specialized receptors expressed by immune effector cells IgG1, 3 - macrophages and neutrophils IgE mast cells, basophils, activated eosinophils (Fc ) Fc of Ag:Ab complexes initiate complement activation Fc can deliver Ab to places would not reach without active transport mucus, tears, milk (IgA) and fetal blood (IgG)

8 Structural variation in Ig constant regions Mature naïve B cells express both IgM and IgD at their surface IgM is the first Ig isotype to be expressed during B cell development IgD is coexpressed with IgM on all mature B cells Immature B cells make mostly transcript while mature B cells make mostly along with transcript When B cell is activated it ceases to coexpress IgD with IgM, either because and sequences have been removed as a consequence of isotype switch or in IgMsecreting plasma cells, because transcription no longer extends through C exons

9 The expression of IgM versus IgD is regulated by RNA processing events operating on a primary RNA transcript that includes both the C and the C exons

10 Structural variation in Ig constant regions Transmembrane and secreted forms of Ig are generated from alternative heavy-chain transcripts All B cells initially express transmembrane (TM) form of IgM After stimulation by Ag, some of their progeny differentiate into plasma cells producing secreted (SC) form of IgM, whereas others undergo isotype switching to express TM Ig of different isotype followed by production of SC Ab of new isotype

11 Structural variation in Ig constant regions Transmembrane and secreted forms of Ig are generated from alternative heavy-chain transcripts In membrane bound form, Ig heavy chain has hydrophobic TM domain of 25aa residues at C-terminus whereas SC form of Ig has hydrophilic C-terminus Two different C-terminus of TM and SC forms of Ig heavy chains are encoded in separate exon and production of two forms is achieved by alternative splicing

12 Transmembrane & secreted forms of Igs are derived from the same H chain sequence by alternative RNA processing

13 Transmembrane & secreted forms of Igs are derived from the same H chain sequence by alternative RNA processing

14 Structural variation in Ig constant regions IgM and IgA can form polymers IgM and IgA C regions include a tailpiece of 18 aa that contains a cysteine residue essential for polymerization J chain promotes polymerization by linking to cysteine of tailpiece In case of IgA, polymerization is required for transport through epithelia Polymerization of Ig is thought to be important in binding of Ab to repetitive epitopes; increasing avidity

15 IgM & IgA immunoglobulins can form multimers via disulfide bonds formed between Fc regions of Ig monomers & association with the non-ig J chain

16 Secondary diversification of Ab repertoire Primary diversification of Ab V(D)J recombination Secondary diversification of Ab - antigen challenge - 3 mechanisms 1) somatic hypermutation 2) gene conversion 3) class switching (isotype switching) - initiated by enzyme activation-induced cytidine deaminase (AID), which is expressed specifically in B cells

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18 Secondary diversification of Ab repertoire Activation-induced cytidine deaminase introduces mutations in genes transcribed in B cells a lack of AID expression a lack of somatic hypermutation and class switching AID deaminates cytidine residues in Ig genes to uridine Additional ubiquitous DNA repair enzymes cooperate with AID to alter single-stranded DNA sequence further - Uracil-DNA glycosylase (UNG) removes pyrimidine base - Apurinic/apyrimidinic endonuclease 1 (APE1) excise the rest of residue, thereby introducing a single-strand nick 3 types of alterations lead to somatic hypermutation, gene conversion and class switching

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22 Secondary diversification of Ab repertoire Rearranged V-region genes are further diversified by somatic hypermutation Somatic hypermutation introduces point mutations in V regions at a very high rate and occurs in germinal centers only after mature B cells have been activated by Ag with signals from activated T cells Affinity maturation: B cell expressing mutant Ig that bind Ag better than original one are preferentially selected to mature into Ab-secreting cells Somatic hypermutation of reaaranged V regions does not occur in T cells

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24 Secondary diversification of Ab repertoire In some species, most Ig gene diversification occurs after gene rearrangement In birds, rabbits, cows, pigs, sheep, horses there is little or no germline diversity in VDJ gene segments Gene conversion: V pseudogene exchanges short sequences with expressed rearranged V gene AID is required for gene conversion

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26 Class switching enables the same assembled V H exon to be associated with different C H genes in course of an immune response The first Ag receptors expressed by B cells are IgM and IgD and the first Ab produced in an immune response is always IgM Later in the immune response, the same assembled V region may be expressed in IgG, IgA or IgE Abs class switching Class switching is stimulated in the course of immune response by external signals such as cytokines released by T cells or mitogenic signals delivered by pathogens Class switch recombination is guided by stretches of repetitive DNA, switch regions

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28 Various differences between Ig can be detected by Abs Isotypes Allotypes Idiotypes Structural variation in Ig constant regions

29 Igs can serve as antigens & can elicit antibodies that can be specific for different regions of their structure