This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Measles is the disease with which the phenomenon of virus-induced immunosuppression was discovered: in 1908 von Pirquet observed that the tuberculin skin test response was transiently depressed during the course of acute measles. Morbilliviruses including measles (MV), canine distemper (CDV) and rinderpest are immunosuppressive. The mechanisms underlying this phenomenon are complex, but viral receptor interactions may play a central role: wild-type MV, CDV and rinderpest virus strains preferentially use the immune cell-specific protein SLAM (human, canine or bovine, respectively) as a receptor. In addition, the MV vaccine strain Edmonston enters cells preferentially also through the ubiquitous regulator of complement activation, CD46, and CD46 interactions modify the immune response to MV. Moreover, post-entry host control evasion mechanisms elicited by the MV non-structural proteins V and C interfere with STAT protein phosphorylation and interferon activation. We will test two hypotheses: first, that SLAM-dependant entry is of central importance for immunosuppression by morbilliviruses. Second, that the V and C proteins favor virus dissemination in immune cells and systemically. Two animal models will be used: macaques for measles and ferrets for canine distemper. We have produced selectively receptor-blind recombinant MVs and CDVs. We are constructing wild type-derived MVs and CDVs in which the expression of V or C, or of both proteins, is silenced or enhanced. Macaques or ferrets will be infected intranasally and the cell types supporting MV and CDV dissemination in PBMC, and in lymphatic and non-lymphatic organs, will be identified. Virulence and immunosuppression will be characterized based on graded parameters including disease signs, leukocyte number, strength and duration of viremia, in vitro lymphocyte proliferation levels, neutralizing antibody liters, and cytokine profile. We predict differential changes in these parameters following infections with viruses defective at the receptor recognition or post-entry level. Results will be interpreted in the context of these predicted outcomes. Candidate mutations for reversion to virulence will be sought in viruses replicating at late disease stages based on functional assays, sequencing, and back-transfer in infectious cDNAs. These experiments will define the relative importance of cell entry through specific receptors and of post-entry host control evasion mechanisms for morbillivirus-induced immunosuppression in two biologically relevant animal systems.
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