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Bet v 1 is the major allergen in birch pollen to which up to 95% of patients sensitized to birch respond. As a member of the pathogenesis-related PR 10 family, its natural function is implicated in plant defense, with a member of the PR10 family being reported to be upregulated under iron deficiency. As such, we assessed the function of Bet v 1 to sequester iron and its immunomodulatory properties on human immune cells. Binding of Bet v 1 to iron quercetin complexes FeQ2 was determined in docking calculations and by spectroscopy. Serum IgE-binding to Bet v 1 with (holoBet v1) and without ligands (apoBet v 1) were assessed by ELISA, blocking experiments and Western Blot. Crosslinking-capacity of apo/holoBet v 1 were assessed on human mast cells and Arylhydrocarbon receptor (AhR) activation with the human reporter cellline AZ-AHR. Human PBMCs were stimulated and assessed for labile iron and phenotypic changes by flow cytometry. Bet v 1 bound to FeQ2 strongly with calculated Kd values of 1 nm surpassing affinities to quercetin alone nearly by a factor of 1000. Binding to FeQ2 masked IgE epitopes and decreased IgE binding up to 80% and impaired degranulation of sensitized human mast cells. Bet v 1 facilitated the shuttling of quercetin, which activated the anti-inflammatory AhR pathway and increased the labile iron pool of human monocytic cells. The increase of labile iron was associated with an anti-inflammatory phenotype in CD14+monocytes and downregulation of HLADR. To summarize, we reveal for the first time that FeQ2 binding reduces the allergenicity of Bet v 1 due to ligand masking, but also actively contributes anti-inflammatory stimuli to human monocytes, thereby fostering tolerance. Nourishing immune cells with complex iron may thus represent a promising antigen-independent immunotherapeutic approach to improve efficacy in allergen immunotherapy. [ABSTRACT FROM AUTHOR]

Based on our I in silico i structural analyses, the IgE-specific effects may be due to epitope masking by the VD3 ligand of two previously described IgE-binding B-cell epitope regions in the molecular surface of Bet v 1 (Figure 1C). Keywords: Bet v 1; birch pollen allergy; immunomodulation; lipocalin-like; vitamin D EN Bet v 1 birch pollen allergy immunomodulation lipocalin-like vitamin D 3801 3804 4 12/01/21 20211201 NES 211201 Abbreviations ANS 1-anilino-8-naphthalene sulfonate BCA bicinchoninic acid BP birch pollen E aff affinity energy FCS foetal calf serum IL interleukin K D binding constant kDa kilodalton MFI mean fluorescence intensity OD optical density PBMCs peripheral blood mononuclear cells PDB protein data bank RA retinoic acid RFUs relative fluorescence units VD3 Vitamin D3 VDRE vitamin D response element ACKNOWLEDGEMENTS We thank Bart R. Blokhuis for excellent technical assistance. [Extracted from the article]

Retinoic acid-loading of the major birch pollen allergen Bet v 1 may improve specific allergen immunotherapy: In silico, in vitro and in vivo data in BALB/c mice Even if this was not due to altered endolysosomal stability of Bet v 1a due to RA binding (Figure S3), it seemed so far like RA-loading transforms the hyperallergen Bet v 1a to a hypoallergen with improved tolerogenic capacity of potential implications for allergen immunotherapy. Mice were first made allergic against the major birch pollen allergen Bet v 1a and subsequently treated intranasally with either the I apo i -Bet v 1a, or with the RA-loaded I holo i -Bet v 1a, or as a control with RA alone. [Extracted from the article]