Dendritic Cells (DCs) play key roles in the initiation of innate and adaptive immune responses during natural infections and vaccination. Two major subsets of DCs, plasmacytoid and conventional DCs, have been described in humans and mice, and exert unique and overlapping immune functions. Among viruses that target DCs, Arenaviruses are of great interest as lethal human pathogens and considered category A agents by NIAID. Importantly, arenavirus infection and subversion of DCs is fundamental for their ability to suppress the immune system, outcompete the infected host and cause disease, but the molecular bases of this process remain poorly understood. The arenavirus nucleoprotein (NP) participates in viral replication, transcription, assembly and suppression of type I interferons (IFN-I), a group of important anti-viral cytokines. We found that plasmacytoid and conventional DCs are productively infected by the prototypic arenavirus LCMV Clone 13 during in vivo murine infection. Furthermore, IFN-I production is inhibited in the vast majority of infected DCs. We hypothesize that the aforementioned functions of arenavirus NPs are achieved by hijacking key host proteins that ultimately enable a successful viral life cycle in DCs and/or DC functional impairment. Our goal is to identify the DC proteins that interact with arenavirus NP and contribute to viral growth and/or immune-evasion. For that, we will first identify NP binding partners in LCMV infected plasmacytoid and conventional DCs by Mass Spectrometry and then validate these physical interactions with other arenavirus NPs. Next, we will knockdown the NP interacting proteins to investigate their importance for arenavirus replication within DCs as well as their role in virus-induced DC functional impairment. These experiments will be performed during the R21 phase and are expected to provide a short-list of host factors that are exploited by arenaviruses to enable their productive replication in DCs and/or disable DC functions in vitro. During the R33 phase of this project, mice or hamsters with genetic alteration of selected NP interacting proteins will be infected with LCMV Clone 13 (infects DCs and suppresses the immune system) or Pichinde arenavirus (causes severe hemorrhagic fever in hamsters), respectively. Viral growth, immune responses and survival will be monitored. These studies will assess the likelihood of targeting selected DC proteins to change the course of an arenavirus infection in vivo. The short-term impact of our studies will be the identification of molecular determinants that play biologically meaningful roles in Arenavirus replication and/or subversion of DCs and that can be manipulated to change the outcome of the infection in small animal models. The long-term impact of our studies will be to translate this new knowledge into novel therapeutic modalities in human infections with arenaviruses and may be other DC-tropic viruses.