Mechanisms of "immune paralysis" caused by filoviruses

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Infections with filoviruses Ebola (EBOV) and Marburg (MARV), lead to the "immune paralysis" characterized by a deficient T cell response and lymphopenia, despite the lack of infection of T cells, and a severe hemorrhagic fever with up to 80% mortality. Over the past decade, researchers identified multiple mechanisms of type I interferon (IFN-I) antagonism mediated by filovirus proteins. However, the mechanisms of the rapid and deleterious disabling of the adaptive immune response by filoviruses remain unknown. Dendritic cells (DC), which are the professional antigen-presenting cells, are believed to be readily infected, but do not undergo normal maturation. Our preliminary data have demonstrated the strong connection between IFN-I antagonism caused by EBOV and the lack of DC maturation. The central hypothesis of the proposal is that the unusually strong and redundant antagonism of the IFN-I response mediated by filovirus proteins and by multiple mechanisms, blocks maturation of DC, resulting in the deficient and/or aberrant stimulation of T cells. The hypothesis will be tested by pursuing three specific aims: 1) Determine the role of IFN-antagonizing domains and IFN-I signaling in the maturation of DC and in the activation, proliferation and apoptosis of T cells; 2) Determine the role of soluble factors secreted by the infected DC and specific inhibitory and pro-apoptotic molecules in the activation, proliferation and apoptosis of T cells; 3) Determine whether the suppression of T cell activation and proliferation or T cell apoptosis during filovirus infection can be reversed by induction of DC maturation. The project will involve the infection of DC derived from donor blood with a recombinant EBOV, its mutants with the IFN-I antagonizing domains disabled, or MARV; stimulation of primary human CD4+ and CD8+ T cells; characterization of the effects of filovirus infection and IFN-antagonizing domains on transcriptomes of infected DC and stimulated T cells by deep sequencing; blockade of IFN-I signaling and immuno-suppressive and pro-apoptotic pathways; and attempts to rescue maturation of infected DC with cytokine cocktails. Together, these experiments represent a broad, integrated approach to investigate the mechanisms of disabling the adaptive immune response by filoviruses.