PROJECT SUMMARY / ABSTRACTViruses commandeer host membrane trafficking to promote viral replication. While this is a critical control pointfor virus infection how viruses hijack host proteins and pathways to promote infection is incompletelyunderstood. Viral-mediated alterations in membrane trafficking are best understood for building replicationcompartments and sites of assembly for RNA viruses. However much less is known about how DNA virusesinfluence membrane remodeling in the host. Human cytomegalovirus (CMV) is a complex DNA virus and amember of the herpesvirus family that establishes a lifelong infection in the host. During infection CMVinduces alterations in membrane trafficking that results in increased biogenesis of multivesicular bodies(MVBs) organelles important for signaling and exocytosis or degradation of cargo. During infection MVBsbecome loaded with virus particles and dense bodies (vesicles of viral tegument protein). The mechanisms bywhich CMV controls host membrane trafficking pathways and particularly MVB biogenesis are notunderstood. Therefore defining the viral proteins and host targets important for the regulation of membranetrafficking is an important goal with important implications for host and virus biology. We have identified twoCMV proteins pUL135 and pUL136 which regulate MVB biogenesis. Recombinant viruses lacking UL135 orUL136 genes exhibit profound alterations in MVB biogenesis and the incorporation of viral cargo into MVB.Consistent with these phenotypes we have identified host interacting proteins for both pUL135 and pUL136proteins that are important for membrane trafficking and MVB biogenesis. The discovery of two viral proteinsand their host interactions strongly position us to define the mechanisms by which CMV modulates membranetrafficking and MVB biogenesis. We hypothesize that UL135- and UL136-host interactions co-opt hostmembrane trafficking and MVB pathways to modulate virus replication. Three aims are proposed to addressthis hypothesis. We will define the association of UL135 and UL136 with subcellular membrane compartmentsand host proteins in Aim 1. In Aim 2 we will define the significance of host interactions to virus replicationmembrane trafficking and incorporation of cargo into MVBs to understand the mechanistic basis of CMV-mediated control of host trafficking. Finally we will investigate the role of UL135 and UL136 and their hostinteracting partners in regulating MVB biogenesis and determine how this regulation impacts viral egress. Ourstudies will define the virus-host interactions and trafficking pathways targeted by CMV and their significance toinfection. Furthermore we anticipate that our findings will uncover common strategies used by many viruses tomanipulate trafficking pathways.