The Blomia tropicalis allergen Blo t 7 stimulates innate immune signalling pathways through TLR 2

Summary Background Although the house dust mite species Blomia tropicalis is a leading cause of allergic diseases in tropical and subtropical regions, the identification and characterization of the allergenic proteins remain incomplete. Objective We aimed to characterize a recombinant form of Blo t 7 ( rB lo t 7) in terms of IgE reactivity, lipid‐binding activity and ability to stimulate innate immunity. Methods The mature Blo t 7 cDNA was cloned by PCR methods for the expression of a secreted form of the allergen in P. pastoris . The IgE reactivity to purified rB lo t 7 as well as the potential cross‐reactivity with Der p 7 was determined by ELISA . The lipid‐binding capacity of rB lo t 7 was assayed using fluorescent lipid probes. The stimulation of TLR 2 signalling pathway by rB lo t 7 was examined in cell activation and reporter assays. Results The amplified mature Blo t 7 cDNA revealed the presence of a 60 base pair insertion compared with the reference sequence registered in the GenBank database. Multiple protein sequence alignments of group 7 mite allergens confirmed that this longer deduced amino acid sequence was the authentic Blo t 7 polypeptide chain. Analysis of IgE reactivity can classify rB lo t 7 as an intermediate B. tropicalis allergen which displayed weak cross‐reactivity with Der p 7. Purified rB lo t 7 was shown to bind selectively the naturally fluorescent lipid probe cis‐parinaric ( cPNA ) with a dissociation constant of 2 μmol/L. The group 7 Blomia allergen stimulated the TLR 2‐, NF ‐ kB ‐ and MAPK ‐dependent production of IL ‐8 and GM ‐ CSF in respiratory epithelial cells. Conclusions & Clinical Relevance Through its propensity to transport fatty acids/lipids and to stimulate TLR 2 signalling pathways in airway epithelial cells, Blo t 7 can represent a key allergen for the initiation of the B. tropicalis ‐induced airway inflammation.