Abstract: Dengue is a rapidly emerging disease that is now endemic in at least 100 countries in Asia, the Pacific, the Americas, Africa, and the Caribbean. Annually it causes an estimated 100 million cases of dengue fever, in which symptoms range from a mild fever to severe dengue hemorrhagic fever in approximately 250.000 cases. Dengue is a major global health problem because of the expanding geographical distribution of both the virus and its mosquito vector, the increased frequency of epidemics, the co-circulation of multiple serotypes and the emergence of severely debilitating dengue hemorrhagic fever in new areas. Currently there is no approved vaccine or antiviral treatment available for dengue viral infection. A detailed understanding of the host functions that determine dengue virus replication and spread may accelerate the development of antiviral drugs. We have recently developed haploid genetic screens in human cells as an efficient approach to perform loss-of-function studies. Using insertional mutagenesis in a haploid human cell line, we have created cell libraries with mutations in most genes. We have shown that this approach can be used to identify host genes required for different pathogens including Ebola virus and Clostridium difficile. In this proposal we aim to find critical host factors that dengue virus uses to establish a successful infection. We will systematically interrogate the function of the genes involved in host antiviral defenses, and we will explore whether these genetic factors can predict the remarkable diversity in symptom severity of infected individuals. We expect these studies will uncover molecular mechanisms of how dengue virus manipulates host cell functions for its replication and spread and how it escapes innate immune responses. Public Health Relevance: Dengue is a mosquito-borne infection that in recent decades has become a major international public health concern due to the severity of its symptoms and the expanding geographical spread of the virus. Detailed mechanistic insight in virus-host interactions will enable the design of specific antiviral therapies and will advance our knowledge of dengue virus pathogenesis.