The goal of this research is to gain new insights in how eosinophils impact asthma and other eosinophil- associated diseases and processes by describing the proteomes of non-activated and acutely and chronically activated human eosinophils. Eosinophils are exceptional in a number of ways: (1) granule content, (2) complement of receptors and other molecules that control activation and trafficking, (3) complement of mediator-generating enzymes, and (4) polarization upon activation by cytokines into a granular compartment and nucleopod, a specialized uropod occupied by the eosinophil's distinctive bilobed nucleus and capped by a signalosome. Deep proteomic analysis will lead to increased understanding of cell-wide processes such as polarization and granule release, generate hypotheses about pathways that are altered in disease, and inform targeted studies. Such studies are feasible. Non-activated eosinophils can be purified reproducibly from blood, and activated acutely with agonists such as interleukin-5 (IL5). Eosinophils purified from broncho-alveolar lavage fluid obtained after segmental antigen challenge provide a unique opportunity to characterize chronically activated cells that have entered and persisted in a tissue milieu enriched in multiple mediators of eosinophilic inflammation. We will identify and quantify the proteins of non-activated and acutely and chronically activated eosinophils, pinpoint sites of phosphorylation, and perform microscopic immuno- localization and other correlative studies to validate the proteomic results and pursue leads that are suggested by the data. Initiatives are proposed based on preliminary results to extend the observation that the signalosome forming in eosinophils activated with IL5 shares components with the well-studied signalosome of B-cells activated via the B-cell receptor; characterize eosinophil Nance-Horan syndrome-like protein 2 (NHSL2), which in preliminary studies was among the proteins most heavily phosphorylated after acute IL5 stimulation; and characterize and localize eosinophil IL18, which was found in unexpectedly high abundance. Understanding dynamics of proteins and post-translational modifications during eosinophil activation is without question necessary to developing a systematic understanding of how human eosinophils arrest in, infiltrate, and perturb tissues in asthma and other diseases. This work will lead to identification of cellular targets with which to modulate eosinophil activation in ways that control inflammation and lead to healing. Going forward, the proteomic techniques that we will establish to accomplish the specific aims can be used to study differences in protein content of eosinophils between normal and individuals with eosinophil-associated diseases and responsiveness of eosinophils to agonists and antagonists of activation.
Disruption or irritation of a barrier presents our immune system with the task of customizing the inflammatory response to fit the threat. Eosinophils, white cell that are easily identified in a blood smear by their abundant plump granules and bilobed nucleus, are a dominant cell in the response mediated by Th2 helper cell population of lymphocytes. This response, although more benign and indolent than the Th1 response, is associated with a considerable disease burden in the form of conditions such as asthma and eosinophilic esophagitis. The contributions of eosinophils to these conditions are incompletely understood. We propose to use cutting-edge global proteomic techniques to assemble comprehensive ?score-cards? of the proteins in eosinophils isolated from blood or inflamed tissues and learn how the proteins are altered acutely when eosinophils are stimulated to enter inflamed tissues. These data will identify cellular targets with which to modulate eosinophils in ways that control eosinophilic inflammation and associated diseases.
| Turton, Keren B; Wilkerson, Emily M; Hebert, Alex S et al. (2018) Expression of novel ""LOCGEF"" isoforms of ARHGEF18 in eosinophils. J Leukoc Biol 104:135-145 |
| Esnault, Stephane; Hebert, Alexander S; Jarjour, Nizar N et al. (2018) Proteomic and Phosphoproteomic Changes Induced by Prolonged Activation of Human Eosinophils with IL-3. J Proteome Res 17:2102-2111 |
| Bortnov, Valeriu; Annis, Douglas S; Fogerty, Frances J et al. (2018) Myeloid-derived growth factor is a resident endoplasmic reticulum protein. J Biol Chem 293:13166-13175 |
| Mosher, Deane F; Wilkerson, Emily M; Turton, Keren B et al. (2017) Proteomics of Eosinophil Activation. Front Med (Lausanne) 4:159 |
| Wilkerson, Emily M; Johansson, Mats W; Hebert, Alexander S et al. (2016) The Peripheral Blood Eosinophil Proteome. J Proteome Res 15:1524-33 |
