Critical Reviews in Immunology. 2016; 36(1): 13-56
Antigen Receptor–Intrinsic Non-Self: The Key to Understanding Regulatory Lymphocyte–Mediated Idiotypic Control of Adaptive Immune Responses
Biochemical Institute of the Medical Faculty of the Christian‑Albrechts‑University, 24098 Kiel, Germany
The clone-specific or idiotypic characters of B as well as T cell antigen receptors (BCRs/TCRs) are associated with (1) the third-complementarity–determining regions (CDR3s) that are created during V(D)J recombination (they scarcely occur in antibody light chains) and (2) BCR idiotopes created by somatic hypermutations (SHMs) during immune responses. Therefore, BCR/TCR idiotypic sites are antigen receptor–intrinsic Non-Self (AgR-iNS) portions that fulfill two tasks: serving as a crucial component of the epitope-binding paratope and serving as target sites for anti-idiotypic BCR/TCR paratopes of other anti-Non-Self clones that are contained in both normal repertoires. The antigen-induced immune response is thus directed not only toward the environmental stimulus but also against the AgR-iNS portions of the directly and further activated clones that form a subsiding idiotypic cascade. These idiotypic chain reactions form a completely integrated idiotypic control circuit among B and T cells which contains all regulatory T and B cells. However, this circuit cannot be viewed as a network of fixed interacting nodes but rather uses the genetic Self as reference. Hence, AgR-iNS offers a mechanistic understanding of regulatory lymphocyte–mediated idiotypic control of adaptive immune responses and reconciles clonal selection and idiotypic network theories hitherto believed to be incompatible.
- PMID: 27480901
- DOI: 10.1615/CritRevImmunol.2016016606
The regulation of cellular activities in the immune system is brought about by a limited number of intrinsic receptors and their corresponding ligands (cytokines, interleukins etc.). Most of these cellular receptors just as their corresponding soluble mediators are fully encoded in the genome. Particular exceptions of this rule are antigen receptors of the adaptive immune system that is destined and indeed able to recognize a seemingly unlimited number not only of environmental but also of synthetic nonnatural antigens. It is obvious that this task cannot be performed with a limited set of genomically encoded receptors. Thus, how can the immune system cope with this challenge? The generally accepted explanation is seen in the recombination of gene segments that together code for the variable domains of antigen-receptors of B (BCR) as well as both types of T cells exhibiting TCRαβ or TCRγδ receptors. All three are assembled from multiple V gene segments, diversity enhancing D gene segments and J gene segments that join the variable portion with constant gene segments (1, 2). While variable regions of antibody light chains (VL), TCRα and TCGγ chains are assembled from V and J gene segments, those of antibody heavy chains (VH), TCRβ and TCRδ chains are assembled from V, D and J gene segments. This V(D)J recombination allows already for huge combinatorial BCR and TCR repertoires both of which, in addition, are greatly enhanced by inaccuracies during the recombination process. These inaccuracies are somatically generated in three different ways.
1.) During recombination of V(D)J segments the recombination-activating genes RAG-1 and RAG-2 generate DNA hairpins which may be opened asymmetrically. Thereby, a few nucleotides of the coding strand plus their complementary nucleotides from the reverse strand form a single-stranded tail. Filling in of the second strand generates a palindromic sequence. In this way a few nucleotides (P nucleotides) of the reverse strand are transferred to the coding sequence where they alter the information of the respective segment (1, 3).
2.) A quantitatively much more important modification at both recombination sites (V-D and D-J) of the BCR heavy chain (but only insignificantly at the light chain) as well as all four polypeptide chains of both types of TCR is created by the enzyme terminal deoxynucleotidyl transferase (TdT) that inserts non-templated nucleotides (N nucleotides) at a variable ratio (1, 4). It has been estimated that TdT function is at least responsible for 90% of the TCRαβ repertoire (5). In contrast to the BCR VL region, TCRα and TCRγ chains that are built by V-J recombination also contain high numbers of N nucleotides (2).
3.) Another important modification during VDJ recombination of BCR and TCR is introduced by exonuclease-mediated deletion of nucleotides from the 3´-end of V gene segments, both ends of the D gene segments and the 5´-end of the J gene segments (1, 2, 5).
Hence, while the first two complementarity-determining regions (CDR1 and CDR2) of all BCR and TCR chains are fully encoded in the genome, the coding sequences of CDR3s of B as well as T cell antigen receptors are somatically generated. This is exemplarily shown in Fig. 1 for the VDJ recombination coding for the variable domain of the immunoglobulin heavy chain. The genomic gene segments are modified by nucleotide deletion and addition of P and N nucleotides to such an extent that the final products of CDR3s are specifically created for each B cell clone. A quantitative impression of these inaccuracies is depicted in Fig. 2 showing nucleotide deletions and additions of monoclonal antibodies reacting with the hapten 2-phenyl-oxazolone (phOx). The main results are: (a) None of the antibodies used full genomic VDJ gene segments. (b) At both ends of the D segment up to 21 of 26 nucleotides could be deleted (Fig. 2C and D). (c) What is even more remarkable, in 16 mAb (36%) the D sequence was deleted to such an extent that it could not be detected. (d) Five mAb (11%) used the full reverse DH gene segment or partial sequences thereof. These and other results highly suggest that CDR3s of B cells of the adaptive immune system in general belong to the Non-Self and do not have an inheritable genomic counterpart.
This is of fundamental importance for two reasons:
1.) Antigen-activated B and T cells not only present peptides of the antigen on MHC molecules but also peptides of their own intrinsic antigen-receptors which can be recognized by other T cell clones contained in the normal repertoire.
2.) Multiple experimental investigations as well as clinical observations have clearly shown that CDR3-associated Non-Self portions of BCR and TCR represent the clone-specific or idiotypic characters of antigen receptors. In addition, new and thereby Non-Self idiotopes are created during thymus-dependent immune responses by somatic hypermutations.
Because of this non-self-nature it is not surprising that the idiotypic characters of BCR and TCR have been observed to occasionally develop strong immunogenicity, in particular during antigen-induced immune responses that are followed by stepwise anti-idiotypic responses that form a subsiding idiotypic chain reaction. Thus, Jerne’s notion “that the immune system of a single animal (or human), after producing specific antibodies to an antigen (or antibody), continues to produce antibodies to the idiotopes of the antibodies that it has itself made” (6) is still valid.
Figure 1. VDJ gene segments coding the antibody heavy chin variable region and modifications creating the third complementarity-determining region CDR-H3 of antibodies
CDR-H1 and CDR-H2 are fully encoded in the genomic VH gene segment. In contrast, CDR-H3 coding sequences are somatically created during recombination of V, D and J gene segments. During this process, all three segments can be modified by deletion of nucleotides through exonuclease activity, addition of P nucleotides and insertions of N nucleotides at the two junctions by terminal deoxynucleotidyl transferase (TdT). Hence, the D-region in the middle of CDR-H3 generally does not conform to the genomic sequences and it is flanked by non-genetic N/P sequences that do not have an inheritable genomic counterpart.
The recognition that the clonotypic or idiotypic characters of B as well as T cell antigen receptors are of Non-Self nature and thus belong to the world of environmental antigens has far reaching consequences for an understanding of multiple functions of the immune system.
- It provides a plausible explanation for the multiple observations of B and T cell idiotypic chain reactions in succession of antigen-induced immune responses and thus reconciles clonal selection and idiotypic network theory.
- The recognition of intrinsic Non-Self portion as idiotypic characters proves that idiotypic regulation turns out to be a physiological necessity.
- This requires reconsideration of the content of immunological memory which is obviously not restricted to antigen-specific B and T cell clones but comprises a net of many idiotypically interconnected clones.
- It explains why antibodies not only exert secondary effector functions of humoral immunity but also actively stimulate regulatory inductor functions. This is of particular importance for the transfer of the immunological experience of the mother to the newborn that aids the initial development of the nascent immune system and even induces long-term effects as, for instance, suppression of IgE responsiveness by allergen-reactive maternal IgG antibodies (idiotypes) or their corresponding anti-idiotypes.
- It explains the immunogenicity of fully human therapeutic antibodies.
- It explains the frequent production of autoantibody-suppressing anti-idiotypic antibodies in healthy individuals.
- It explains the old observation that thymus-dependent immune responses provoke the production of immunoglobulins that, surprisingly and in contrast to the clonal selection theory, do not react with the antigen (7). It is striking that these antigen-non-specific responses are much stronger than the specific antibody production.
- As part of the function of regulatory B and T cells interclonal idiotypic regulations among B and T cells are clearly visible during T cell vaccination (TCV) for the treatment of autoimmune and malignant diseases.
In summary, on the basis of the combinatorial repertoire the somatic creation of clone-specific BCR and TCR Non-Self portions generates the junctional repertoire allowing the immune system to respond to a seemingly unlimited number of antigens. The intrinsic Non-Self portions of antigen-activated B and T cells induce further anti-idiotypic responses that form a subsiding idiotypic chain reaction. These reactions seem to represent a primary regulatory principle that initiates and regulates the differentiation of lymphocytes during antigen/autoantigen-induced immune responses before the effector functions are executed by cytokines.
Figure 2. Modification of VDJ gene segments in anti-hapten antibodies
D-altered ΔD-iD mice containing a single modified DFL16.1 D gene segment with a length of 26 base pairs in their genome (8) were immunized with the hapten 2-phenxl-oxazolone coupled to chicken serum albumin. Monoclonal antibodies (n = 44) were prepared during various stages of primary and memory responses and V region sequences were determined (9). Twenty three mAb (~52%) used modified DH gene sequences and five mAb (~11%) reverse DH sequences. Nucleotide deletion and addition of N/P nucleotides is shown for these two groups of antibodies: A – nucleotide deletion at 3’-VH, B – addition of N/P nucleotides at the VH-DH junction, C – nucleotide deletion at 5’-DH, D – nucleotide deletion at 3’-DH, E – addition of N/P nucleotides at the DH-JH junction, F – deletion of nucleotides at 5’-JH. The medium values of deletion and addition are indicated by a broken red line with a red triangle on top. In 16 additional mAb (~36%) the DH coding sequence is crippled by exonuclease activity to such an extent that it could not be detected.
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Acknowledgements: This work was supported by personal grants from the Deutsche Forschungsgemeinschaft and SFB 877.
Hilmar Lemke, Ph.D.
Biochemical Institute of the Medical Faculty of the Christian‑Albrechts‑University at Kiel, D‑24098 Kiel, Germany