Genetic variability of the major birch pollen allergen Bet v 1 and characterization of B- and T-cell epitopes

The hanging birch pollen collected in the period April – May 2020, 2021 in the territory of six regions of the Republic of Belarus was studied. Recombinant plasmid DNA was obtained. A nucleotide sequence of cloned fragments was determined. The results on the spectrum of isoforms of the Bet v 1 protein were analyzed. The obtained sequences corresponded to one degree or another to 11 genetic variants of the studied allergen. There were 7 isoforms of Bet v 1 defined within one tree. The predominant isoform of the birch pollen allergen Bet v 1 in the territory of the Republic of Belarus was Bet v 1.0101 (Bet v 1a, X15877.1). The identified variants were analyzed for their potential allergenicity by screening amino acids that according to the literature data were identified as affecting IgE-binding. The analysis of amino acid residues included in the IgE-binding conformational epitopes revealed amino acid substitutions exhibiting the multidirectional (high or low) IgE-binding activity in positions 31, 58, 113, 114, 126. The structure of dominant epitopes recognized by the T-cell receptor was studied. It was found that the C-terminal immunodominant T-cell epitope Bet v 1_143–157 is highly conserved among various isoforms of the allergen in contrast to the epitope Bet v 1_78–93 located in the central region. The revealed amino acid substitutions of the studied sites can affect the activation of T-cells, cross-reactivity and significantly increase the variability of the expected IgE-mediated reaction.

1. Breiteneder H., Kraft D. The History and Science of the Major Birch Pollen Allergen Bet v 1. Biomolecules, 2023, vol. 13, no. 7, art. 1151. https://doi.org/10.3390/biom13071151

2. von Loetzen C. S., Jacob T., Hartl-Spiegelhauer O., Vogel L., Schiller D., Spörlein-Güttler C., Schobert R., Vieths S., Hartl M. J., Rösch P. Ligand recognition of the major birch pollen allergen Bet v 1 is isoform dependent. PLoS One, 2015, vol. 10, no. 6, art. e0128677. https://doi.org/10.1371/journal.pone.0128677

3. Schenk M. F., Gilissen L. J., Esselink G. D., Smulders M. J. Seven different genes encode a diverse mixture of isoforms of Bet v 1, the major birch pollen allergen. BMC Genomics, 2006, vol. 7, art. 168. https://doi.org/10.1186/1471-2164-7-168

4. Wagner S., Radauer C., Bublin M., Hoffmann-Sommergruber K., Kopp T., Greisenegger E. K., Vogel L., Vieths S., Scheiner O., Breiteneder H. Naturally occurring hypoallergenic Bet v 1 isoforms fail to induce IgE responses in individuals with birch pollen allergy. Journal of Allergy and Clinical Immunology, 2008, vol. 121, no. 1, pp. 246–252. https://doi.org/10.1016/j.jaci.2007.08.006

5. Ferreira F. D., Hoffmann-Sommergruber K., Breiteneder H., Pettenburger K., Ebner C., Sommergruber W., Steiner R., Bohle B., Sperr W. R., Valent P. Purification and characterization of recombinant Bet v I, the major birch pollen allergen. Immunological equivalence to natural Bet v I. Journal of Biological Chemistry, 1993, vol. 268, no. 26, pp. 19574–19580. https://doi.org/10.1016/s0021-9258(19)36554-8

6. Erler A., Hawranek T., Krückemeier L., Asam C., Egger M., Ferreira F., Briza P. Proteomic profiling of birch (Betula verrucosa) pollen extracts from different origins. Proteomics, 2011, vol. 11, no. 8, pp. 1486–1498. https://doi.org/10.1002/pmic.201000624

7. Swoboda I., Jilek A., Ferreira F., Engel E., Hoffmann-Sommergruber K., Scheiner O., Kraft D., Breiteneder H., Pittenauer E., Schmid E., Vicente O., Heberle-Bors E., Ahorn H., Breitenbach M. Isoforms of Bet v 1, the major birch pollen allergen, analyzed by liquid chromatography, mass spectrometry, and cDNA cloning. Journal of Biological Chemistry, 1995, vol. 270, no. 6, pp. 2607–2613. https://doi.org/10.1074/jbc.270.6.2607

8. Schenk M. F., Cordewener J. H., America A. H., van’t Westende W. P., Smulders M. J., Gilissen L. J. Characterization of PR-10 genes from eight Betula species and detection of Bet v 1 isoforms in birch pollen. BMC Plant Biology, 2009, vol. 9, art. 24. https://doi.org/10.1186/1471-2229-9-24

9. Radauer C., Nandy A., Ferreira F., Goodman R. E., Larsen J. N., Lidholm J., Pomés A., Raulf-Heimsoth M., Rozynek P., Thomas W. R., Breiteneder H. Update of the WHO/IUIS Allergen Nomenclature Database based on analysis of allergen sequences. Allergy, 2014, vol. 69, no. 4, pp. 413–419. https://doi.org/10.1111/all.12348

10. Allergen Nomenclature. Available at: http://www.allergen.org/index.php (accessed 1 April 2024). 11. Bijli K. M., Singh B. P., Sridhara S., Arora N. Isolation of total RNA from pollens. Preparative Biochemistry and Biotechnology, 2001, vol. 31, no. 2, pp. 155–162. https://doi.org/10.1081/PB-100103381

11. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proceedings of the National Academy of Sciences of the United States of America, 1977, vol. 74, no. 12, pp. 5463–5467. https://doi.org/10.1073/pnas.74.12.5463

12. Basic Local Alignment Search. Available at: https://blast.ncbi.nlm.nih.gov/Blast.cgi. (accessed 1 April 2024)

13. Hoffmann-Sommergruber K., Vanek-Krebitz M., Radauer C., Wen J., Ferreira F., Scheiner O., Breiteneder H. Genomic characterization of members of the Bet v 1 family: genes coding for allergens and pathogenesis-related proteins share intron positions. Gene, 1997, vol. 197, no. 1–2, pp. 91–100. https://doi.org/10.1016/S0378-1119(97)00246-1

14. Gajhede M., Osmark P., Poulsen F. M., Ipsen H., Larsen J. N., Joost van Neerven R. J., Schou C., Løwenstein H., Spangfort M. D. X-ray and NMR structure of Bet v 1, the origin of birch pollen allergy. Nature Structural and Molecular Biology, 1996, vol. 3, pp. 1040–1045. https://doi.org/10.1038/nsb1296-1040

15. Lund G., Christensen L. H., Ihlemann J., Andersen P. S., Wambre E., Würtzen P. A., Gupta S. T cells specific to multiple Bet v 1 peptides are highly cross-reactive toward the corresponding peptides from the homologous group of tree pollens. Frontiers in Immunology, 2023, vol. 14. https://doi.org/10.3389/fimmu.2023.1291666

16. Friedl-Hajek R., Radauer C., O’Riordain G., Hoffmann-Sommergruber K., Leberl K., Scheiner O., Breiteneder H. New Bet v 1 isoforms including a naturally occurring truncated form of the protein derived from Austrian birch pollen. Molecular Immunology, 1999, vol. 36, no. 10, pp. 639–645. https://doi.org/10.1016/s0161-5890(99)00078-4