The mechanism by which antigen-presenting cells (APCs) convert free influenza virus to a form recognized by major histocompatibility complex-restricted helper T cells (TH) will be investigated. From the nominal antigen specificities of TH clones arising in response to influenze virus infection or immunization, a number of different TH determinants on each of the major influenze structural proteins, as well as at distinct sites on an individual protein, are known to be expressed on the APC surface as a result of the presentation process. This research will focus on the route and mechanisms by which these viral entigens, including both external glycoproteins and internal virion proteins, are handled and expressed by APCs. A major distinction must be made as to whether a given determinant is introduced as part of the intact virus or as an isolated protein or peptide: the influenza virion possesses a mechanism for infectious cell entry involving 1) binding to the host cell, and 2) fusion into host membranes. The effect of these activities on the handling of antigen and function of the APC will be assessed in the following experiments: A) The relationship of cell-binding activity to the efficiency and rate of presentation of viral determinants will be studied by perturbing binding of virus to APCs with neruaminidase or inhibitory antibodies and by use of peptides and free proteins lacking cytophilic activity. B) The role of intracellular mechanisms in processing of virus will be approached by comparing the effect of different inhibitory treatments (e.g., temperature, lysosomotropic agents, enzyme inhibition) upon presentation of viral antigens C) The effect of virus fusion will be studied both by abrogation of the fusion capability of virus and introduction of viral proteins directly into the cytoplasmic membrane of the APC. D) The expression of TH determinants on the APC surface will also be approached by using monoclonal antibodies specific for short peptides embodying known antigens for antiviral TH in studies of blocking TH recognition and binding to the APC. The major objectives of this research are to characterize APC-mediated events which underlie TH stimulation in viral infections, and the relationship of the antigen presentation process to the selection and properties of the major T cell antigenic sites on viral proteins.
