Diversity in the CDR3 region of V(H) is sufficient for most antibody specificities

All rearranging antigen receptor genes have one or two highly diverse complementarity determining regions (CDRs) among the six that typically form the ligand binding surface. We report here that, in the case of antibodies, diversity at one of these regions, CDR3 of the V(H) domain, is sufficient to permit otherwise identical IgM molecules to distinguish between a variety of hapten and protein antigens. Furthermore, we find that somatic mutation can allow such antibodies to achieve surprisingly high affinities. These results are consistent with a model in which the highly diverse CDR3 loops are the key determinant of specificity in antigen recognition in both T cell receptors (TCR) and antibodies, whereas the germline-encoded CDR1 and CDR2 sequences are much more cross-reactive.     

T cell receptor CDR3 loop length repertoire is determined primarily by features of the V(D)J recombination reaction

The third complementarity-determining region (CDR) of the TCR alpha and beta chains forms loops that engage amino acid residues of peptides complexed with MHC. This interaction is central to the specific discrimination of antigenic-peptide-MHC complexes by the TCR. The TCRbeta chain CDR3 loop is encoded by the Dbeta gene segment and flanking portions of the Vbeta and Jbeta gene segments. The joining of these gene segments is imprecise, leading to significant variability in the TCRbeta chain CDR3 loop length and amino acid composition. In marked contrast to other pairing antigen-receptor chains, the TCR beta and alpha chain CDR3 loop size distributions are relatively narrow and closely matched. Thus, pairing of TCR alpha and beta chains with relatively similar CDR3 loop sizes may be important for generating a functional repertoire of alpha beta TCR. Here we show that the TCRbeta chain CDR3 loop size distribution is minimally impacted by TCRbeta chain or alpha beta TCR selection during thymocyte development. Rather, this distribution is determined primarily at the level of variable-region gene assembly, and is critically dependent on unique features of the V(D)J recombination reaction that ensure Dbeta gene segment utilization.     

Analysis of T cell receptor alpha-chain variable region (Valpha) usage and CDR3alpha of T cells infiltrated into lesions of psoriasis patients

Psoriasis is a common inflammatory skin disease and is considered as T cell-mediated immune response. In this study, we analyzed T cell receptor alpha-chain variable region (TCR Valpha) usage in the lesions of psoriasis patients using 5'-RACE. As the results, Valpha1, -2, -7, -8, -10, -11, -12, and -23 were commonly detected in psoriatic lesions and comparison of expressions of these Valpha types between psoriasis patients and healthy individuals showed that Valpha1, -7, -11, and -12 were highly increased in psoriasis patients than in healthy individuals. Compared with atopy dermatitis patients, the expressions of Valpha1 and Valpha7 were increased in psoriasis patients. Then, to identify CDR3alpha of T cells infiltrated in psoriatic lesions, we examined which type of J gene segment was rearranged with Valpha1 or Valpha7, which the expressions was specifically increased in psoriatic lesions. The result showed that the V-J rearrangements between the examined patients were not equivalent and their frequencies were diverse, however, several common rearrangements such as Valpha1-Jalpha13, -23, -27, or -34 and Valpha7-Jalpha12, -33 were detected. The results in this study might provide the clue to define the characteristics of T cells associated with the pathogenesis of psoriasis.     

Hb J- Meerut [alpha 120 (H3) Ala ->Glu (alpha1)] in a Turkish male

Hb J Meerut is an infrequently found alpha-globin variant. It has previously been reported in various populations around the world. One particular case reported in 1994 included a Turkish family. In this report, details of a second case of Hb J Meerut in a Turkish male who is unrelated to the first family are described. In the present case a slight increase in the oxygen affinity of Hb J Meerut, relative to that of the normal control, has been observed as detected by low p50 values in arterial whole blood. Additionally, a slight increase in red blood cell count, as compared against a normal individual, was observed.     

The amino acid residue at position 95 and the third CDR region in the H chain determine the ceiling affinity and the maturation pathway of an anti-(4-hydroxy-3-nitrophenyl)acetyl antibody

Two groups of anti-(4-hydroxy-3-nitrophenyl)acetyl (NP) Abs each possessing a different amino acid, Tyr or Gly, at position 95, appeared respectively at early and late stages of immunization. The early Abs predominantly harbored Tyr95 and were referred to as the Tyr95 type. These had ～100-fold lower ceiling affinity than the late Abs harboring Gly95, which were referred to as the Gly95 type. We found that in order to raise affinity, the Tyr95 type utilized a mutation at position 33 in V(H), while the Gly95 type used multiple mutations in both V(H) and V(L), and that the effect of the mutations was reciprocal; the former mutation had a positive effect on Tyr95 type Abs but a negative effect on Gly95 type Abs, and vice versa. The reciprocal effect of these mutations on affinity enabled us to assess the type of Abs prepared by introducing 20 different amino acids at position 95. We found that Abs harboring Lys95, Arg95, Pro95, and Tyr95 belonged to the Tyr95 type and those with Ala95 and Gly95, to the Gly95 type. Since this dependency on the amino acid at position 95 was observed in H chains whose third CDR (CDR 3H) consisted of 9 amino acids and not 11, the CDR 3H region was also considered to play an important role in determining the maturation pathway and the magnitude of the ceiling affinity.     

Thymocyte differentiation in γ-irradiated severe-combined immunodeficient mice: characterization of intermediates and products of V(D)J recombination at the T cell receptor α locus

Treatment with DNA-damaging agents promotes rescue of V(D)J recombination, limited thymocyte differentiation, and development of thymic lymphomas in severe-combined immunodeficient (SCID) mice. One intriguing aspect of this system is that irradiation rescues rearrangements at the T cell receptor (TCR) β, γ and δ loci, but not at the TCR α locus. Current models posit that only those loci that are recombinationally active at the time of irradiation can be rescued. Here, we employ sensitive, semiquantitative ligation-mediated polymerase chain reaction assays to detect a specific class of recombination intermediates, hairpin coding ends, at the TCR α locus. We found that Jα-coding ends are undetectable in unirradiated SCID thymocytes, but accumulate after irradiation at times coincident with the emergence of a CD4⁺CD8⁺ thymocyte population. Coding joints produced by joining of these ends, however, are extremely rare. To test whether the presence of hairpin coding ends at TCR α is sufficient for irradiation-mediated rescue of coding joint formation, we administered a second dose of γ-irradiation after abundant CD4⁺ CD8⁺ thymocytes and hairpin TCR α coding ends had accumulated. This treatment failed to stimulate rescue of TCR α coding joints. Thus, the presence of hairpin coding ends at the time of irradiation, while perhaps necessary, is not sufficient for rescue of V(D)J rearrangements. These results support a refined model for irradiation-mediated rescue of TCR rearrangements in SCID mice.     

Reactivity of human anti-alpha-galactosyl IgG antibody with alpha(1-->3)-linked galactosyl epitopes exposed on basement membranes and on glomerular epithelial cells: an in vitro and in vivo study in the mouse

Anti-alpha-galactosyl antibody (a-Gal Ab) is a human natural antibody belonging to the IgG class, found in high titres in all normal sera regardless of blood group, and specifically recognizing alpha (1-->3)-linked galactosyl residues. We have observed by radioimmunoassay, ELISA, passive haemagglutination and immunofluorescence blocking studies that affinity-purified a-Gal Ab reacted with mouse laminin, but not with the other mouse basement membrane proteins tested; it was able to fix complement in vitro. When injected intravenously into mice, the a-Gal Ab was found to mainly accumulate in kidneys, liver, spleen and lungs. No acute respiratory distress syndrome was observed shortly after the i.v. injection of 100 or 200 microg of antibodies. These doses of a-Gal Ab were also unable to induce acute glomerular injury. However, in primary cultures, the a-Gal Ab (100 or 200 microg per ml of medium) was shown to impair the attachment of mouse glomerular epithelial cells to mouse laminin and to elicit complement-dependent cell damage. The data indicate that the a-Gal Ab can interact in vitro and/or in vivo with alpha (1-->3)-linked galactosyl residues exposed on murine laminin or on murine cultured glomerular epithelial cells. Although this antibody fails to be pathogenic when administered at low doses in the intact animal, similar doses can alter some metabolic properties of these cells in vitro.