Isogenic cells respond in a heterogeneous manner to interferon. Using a micropatterning approach combined with high-content imaging and spatial analyses, we characterized how the population context (position of a cell with respect to neighboring cells) of epithelial cells affects their response to interferons. We identified that cells at the edge of cellular colonies are more responsive than cells embedded within colonies. We determined that this spatial heterogeneity in interferon response resulted from the polarized basolateral interferon receptor distribution, making cells located in the center of cellular colonies less responsive to ectopic interferon stimulation. This was conserved across cell lines and primary cells originating from epithelial tissues. Importantly, cells embedded within cellular colonies were not protected from viral infection by apical interferon treatment, demonstrating that the population context-driven heterogeneous response to interferon influences the outcome of viral infection. Our data highlights that the behavior of isolated cells does not directly translate to their behavior in a population, placing the population context as one important factor influencing heterogeneity during interferon response in epithelial cells.Synopsis The heterogeneous response to interferons in epithelial cells is determined by the population context (i.e. a cell’s positions within a population) through cell polarization and basolateral localization of the interferon receptor. Isogenic epithelial cells respond in a heterogeneous manner to type I and type III interferons. The population context (position of a cell within a population) drives the heterogeneous response to interferons, with cells located at the edge of a population being more responsive than cells located in the center of a population. Cells in the center of a population are refractive to interferon treatment from their apical side due to the polarized basolateral distribution of the interferon receptors. By controlling the response to interferon treatment, the population context affects susceptibility to viral infection.The heterogeneous response to interferons in epithelial cells is determined by the population context (i.e. a cell’s positions within a population) through cell polarization and basolateral localization of the interferon receptor.Deutsche Forschungsgemeinschaft (DFG)http://dx.doi.org/10.13039/501100001659Chica and Heinz Schaller Foundation (CHS-Stiftung)http://dx.doi.org/10.13039/501100012284Deutscher Akademischer Austauschdienst (DAAD)http://dx.doi.org/10.13039/501100001655UF | UF Health | College of Medicine, University of Florida (UF College of Medicine)http://dx.doi.org/10.13039/10000827
