T cells play an important role in the pathogenesis of allergic diseases. However, potential immunogens of allergenic T cell responses have traditionally been limited to those that induce IgE responses. Timothy grass (TG) pollen is an inhaled allergen for which major IgE-reactive allergens have also been shown to trigger Th2 responses. Here we examined whether other TG pollen proteins are recognized by Th2 responses independently of IgE-reactivity. A TG pollen extract was analyzed by 2D-gel electrophoresis and IgE/IgG immunoblots using pooled sera from allergic donors. Mass spectrometry in combination with de-novo sequencing of the whole TG pollen transcriptome identified 93 novel proteins for further study, 64 of which were not targeted by IgE. Peptides from the novel TG proteins were screened for T cell reactivity in PBMCs from allergic donors. Strong IL-5 production was detected in response to peptides from several novel proteins, most of which were not targeted by IgE. Responses against the dominant novel epitopes were associated with memory T cells and could be detected directly ex vivo after Th2 cell enrichment. These findings demonstrate that a combined unbiased transcriptomic-, proteomic-, and immunomic approach identifies a greatly broadened repertoire of protein antigens targeted by T cells involved in allergy pathogenesis. The discovery of proteins that induce Th2 cells but are not IgE reactive may allow the development of safer immunotherapeutic strategies.
- Copyright © 2013 by The American Association of Immunologists, Inc.