Immunogenetacs (1995) 42:53-58 © Springer-Verlag 1995 Heimo Breiteneder • Otto Scheiner • Roswitha Hajek Wolfgang Hulla • Robert Hiittinger • Gottfried Fischer Dietrich Kraft • Christof Ebner Diversity of TCRAV and TCRBV sequences used by human T-cell clones specific for a minimal epitope of Bet v 1, the major birch pollen allergen Received: 19 January 1995 / Revised: 9 March 1995 Abstract T-cell clones (TCC) were raised from the pe- ripheral blood of patients suffering from tree pollen allergy. All TCC were restricted by HLA-DR molecules. In order to investigate possible intervention targets in Type I allergic diseases, we examined T-cell receptor (TCR) cc and ~ chain nucleotide sequences of five allergen-reactive human CD4 + TCC specific for a C-terminal epitope (BV 144) of Bet v 1, the major birch pollen allergen. Proliferation assays using synthetic peptides revealed the 10-mer LRAVESYLLA as minimal epitope for three TCC; two TCC also displayed reactivity with the nonapeptide LRAVESYLL. Two TCC expressed TCRBV2S3, all other BV144-specific TCC used diverse TCRAV and TCRBV gene segments_ Moreover, the junctional regions encoding the third complementary determining regions (CDR3) of the TCR showed a striking heterogeneity in length and amino acid composition. Introduction The T-cell antigen receptor (TCR) of mature T lymphocytes recognizes antigens in the form of short peptide fragments bound to major histocompatibility complex (MHC) mole- cules on the surface of antigen presenting cells (APC; Schwartz 1985). Over 90% of peripheral blood T lympho- cytes express the TCR in the form of a disulfide-linked heterodimer composed of a TCRc¢ and TCR~ chain (Haskins et al. 1983; Meuer et al. 1983)_ Each chain consists of a variable region which confers the specificity of the TCR for the peptide/MHC-complex, and a constant region that attaches this variable region to the cell surface. During thymic maturation, unique variable region genes are created by recombination of germ-line encoded variable Nevertheless, all TCC showed an arginine residue in the (V), diversity (D, for the ~ chain only) and joining (J) N-terminal region of their TCRBV CDR3 loops. Therefore, segments. Combinatorial diversity is generated by the therapeutical strategies aimed at the clonal deletion of joining of unique combinations of these segments as well allergen-specific T-cell clones, providing help for IgE synthesis, will not be feasible. Our results cast a doubt on the theory that the CDR3 exclusively provides the primary contact with the peptide bound in the major histocompat- ibility (MHC) groove, and suggest additional interaction with MHC class II. The nucleotide sequence data reported in this paper have been submitted to the EMBL nucleotide sequence database and have been assigned the accession numbers Z47366-Z47376 H. Breiteneder • O. Scheiner • R. Hajek • W. Hulla • R. Htittinger • D. Kraft • C. Ebner (~) Institute of General and Experimental Pathology, AKH-EBO, University of Vienna, W~ihringer Giirtel 18-20, A-1090 Vienna, Austria G. Fischer Institute of Blood Group Serology, AKH, Wghringer Gtirtel 18-20, A-1090 Vienna, Austria as the random association of any cz chain with any [~ chain. Further diversity of the variable domain is contributed by the loss (through nucleotide "nibbling") or template-inde- pendent addition of N-region nucleotides at V-(D)-J junc- tions (Davis and Bjorkman 1988). After rearrangement, thymic selection leads to abortion or rescue from abortion of developing lymphocytes (von Boehmer 1992). Unlike in immunoglobulin (Ig)-rearrangement, somatic hypermuta- tion does not seem to play a significant role in generating TCR diversity (Ikuta et al. 1985). However, the structure of the TCR is similar to that of Ig, containing three or four hypervariable loops that correspond to the complementar- ity-determining regions [(CDR) (Clothia et al. 1988)]. The CDR1 and CDR2 loops of the TCR are encoded by germ- line V-segment sequences and are believed to interact with the oc helices forming the side walls of the binding groove of the MHC molecules. In contrast, the CDR3 loops are encoded by the V-(D)-Jjuctional regions and are believed to provide the primary contact with the peptide bound in the MHC binding groove (Davis and Bjorkman 1988; Clothia et al. 1988).