Qualitative spatiotemporal characterization of collective cell death applied to ferroptosis

While most forms of cell death are studied under the premise that an individual cell’s death has no effect on neighboring cells, emerging evidence indicates that different cell death mechanisms also have distinct effects on cell populations that extend beyond the consequences to individual cells. Such population-scale effects remain largely unexplored, due to lack of computational and imaging-based approaches that will allow for their systematic and quantitative characterization. We developed methods to measure the spatiotemporal dynamics of collective cell death at the single cell level. Specifically, we decoupled two cellular mechanisms that together contribute to the spreading of cell death within a population: (1) autonomous death caused by external stress; and (2) non-autonomous death, the propagation of a death signal from one cell to its neighbors. We applied this methodology to validate that ferroptosis, an iron-dependent form of cell death, was dominated by non-autonomous cell death while apoptosis by autonomous cell death. Precise spatiotemporal death characterization of different forms of ferroptosis revealed (1) “collective” modes of erastin-induced ferroptosis that involve, beyond propagative signals, more cell-autonomous death that together accelerate total death; and (2) The inhibition of the ferroptosis-inhibiting enzyme GPX4 leads to short-time propagative effects on neighboring cells, in contrast to previous description as autonomous death. Both of these population-scale effects occur independently of the final execution stage of cell death involving rupture of cell membrane, suggesting involvement of upstream signaling mechanisms that can coordinate individual cell fates.