The Zika epidemic that began in Brazil and has spread to Central America and US territories, caused by the Asian Zika virus (ZIKV) strain, has been reported by the CDC as definitively linked to severe birth defects – microcephaly, miscarriage and stillbirth. Detection of ZIKV RNA in the placenta and fetus and in maternal circulation for months, calcifications associated with long-standing infection, and intrauterine growth restriction indicate that virus-induced pathology impairs placental functions. To address the issue of transplacental ZIKV transmission, we propose a collaborative project between Dr. Lenore Pereira (UCSF) and Dr. Eva Harris (UC Berkeley), experienced investigators with complementary expertise in congenital infection of the human placenta and Flavivirus research, respectively. Our preliminary studies reveal that prototype and recently iso- lated Nicaraguan ZIKV strains infect cells in placental explants and primary cells isolated from human placenta that express AXL, Tyro3 and TIM1 tyrosine kinase receptors, which mediate ZIKV and the closely related dengue virus (DENV) infection in skin. Infected placental cells, including fetal amniotic epithelial cells, placental fibroblasts and trophoblast progenitor cells, developed cytopathology and expressed ZIKV envelope and non- structural NS3 proteins, and virus titers released depended on the receptors expressed and gestational age. Indicative of infection route, AXL was detected in decidua (uterine decidual cells and invasive cytotrophoblasts), chorionic villi (placental fibroblasts, Hofbauer cells and blood vessels) and fetal membranes (amniotic epithelial cells and trophoblast progenitor cells). Differential expression of receptors suggests how ZIKV could infect the decidua and spread to the placenta, fetus and amnion-chorion membranes. Further, in endemic regions, cross- reactive pre-existing antibodies to DENV could play a critical role in protection or pathogenesis of ZIKV in placenta tissues during pregnancy. Our overarching hypothesis is that placental primary cells and explants of decidua, chorionic villi and amnion-chorion membranes can be used as a model of the human placenta to define molecular mechanisms of ZIKV infection by free virions; the role of antibodies in neutralization, trans- cytosis, and enhancement of ZIKV infection in the placenta; and the translational impact of inhibitors and thera- peutic neutralizing monoclonal antibodies (MAbs). Our Specific Aims are: Aim 1. Identify cells at the uterine- placental interface that are susceptible to infection by Nicaraguan ZIKV strains in primary cells and placental explants, characterize cognate receptors, and identify small-molecule inhibitors of infection. Aim 2. Study the neutralizing and potentially enhancing role of ZIKV-specific and DENV-cross-reactive antibodies on infection and transcytosis in primary placental cells and explants and determine the therapeutic potential of neutralizing MAbs with modified Fc that preclude antibody-dependent enhancement. The proposed studies will reveal molecular mechanisms of ZIKV infection, routes of virus transmission to the fetus, and therapeutic potential for engineered antibodies and small molecule inhibitors that block infection and prevent congenital disease.