H. James Wedner, M.D.

DEPARTMENT OF Int Med - Immunology
Keywords: allergy, IgE, pollen

My laboratory is interested in the immunobiology of allergens with particular emphasis on diagnosis and therapy. Our major thrust is a thorough examination of the allergens contained in pollen proteins and in fungi, both spores and micelia. Specifically, we wish to identify and characterize the allergens in terms of their amino acid sequences, IgE-reactive epitopes and those epitopes that react with CD4⁺ T cells (when presented by self APCs). We have chosen two model systems for this study: 1) the allergens in white oak pollen, the most common cause of tree seasonal pollinosis in the Midwest and 2) allergenic fungi, including Epicoccum nigrum, a mold originally thought to be important only in rural areas but which we recently have demonstrated to be the cause of allergies in almost half of the mold-sensitive individuals in St. Louis, and other indoor and outdoor molds.

Allergic individuals are identified by skin testing or by an enzyme-linked immunoassay (ELISA) for IgE directed against allergens. Sera from sensitive patients is then used to identify individual allergens by immunoblotting following separation of allergenic extracts by single dimension SDS-PAGE or two-dimensional electrophoresis. Using l-D techniques, we have identified at least 23 allergens in white oak pollen and more than 40 allergens in E. nigrum spore and micelial extracts. Some fungal allergens are unique to spore or mycelia, while many are common to both. The 2-D technique increased the number of allergens in white oak to 28. In addition, 2-D electrophoresis has allowed us to perform direct N-terminal amino acid sequencing on allergens identified by immunoblotting. To further characterize the allergens in white oak, a cDNA library has been prepared using mRNA isolated from oak inflorescences in the lZAP expression vector. We have found that the fusion proteins expressed by this library bind oak pollen-specific IgE. This library will be used to characterize the major and minor allergens of oak pollen. Similar studies also are in progress using mRNA isolated from E. nigrum spores and micelia.

The sequencing studies coupled with IgE will allow us to identify the IgE-reactive epitopes of individual allergens. The T cell-reactive areas will be identified using allergen-specific T cell clones which we have derived from allergic individuals. To date, we have established clones from two individuals who are sensitive to white oak pollen. The fine specificity of these clones will be identified using purified allergens and allergenic fragments synthesized from the known amino acid sequences of the allergens. These studies should allow us to differentiate the areas of the stimulation of CD4⁺ T cell and should yield new insights into human allergic diseases and their treatment.