Curated Optogenetic Publication Database

Abstract: Pyroptosis, an inflammatory form of cell death, is associated with large cell swelling and plasma membrane rupture. Recently, such swelling has been shown to occur in a two steps fashion, but the precise molecular and biophysic mechanisms driving the process remain elusive. We demonstrate through advanced quantitative microscopy that, between the two swelling phases, cell volume stabilizes, while plasma membrane permeability to ions and small molecules is markedly elevated due to the formation of pores. From a biophysical perspective, how such a volume plateau exists is puzzling as ion pumps should not regulate the cell volume in these conditions. To address this, we developed a physical model based on an ions pump-and-leak framework, incorporating the dynamics of non-selective pore formation. We experimentally identify two distinct pore permeability dynamics, associated to an increase in the water filtration coefficient and to an ion selectivity decrease due to pore opening. Altogether our results suggest the existence of two mechanistically different pore types, likely driven by separate molecular players. Our findings provide fundamental insights into the biophysics of cell death and may have broader implications for understanding membrane rupture in other pathological contexts. Significance Statement: Among various programmed lytic cell death, pyropytosis is marked by dramatic changes in cell shape and large fluctuations in volume, fundamentally altering the cell’s physical properties. These biophysical changes are not mere byproducts but integral components of the death process, closely interacting with molecular events. By combining optogenetics, quantitative microscopy, and modeling, we show that a progressive increase in plasma membrane permeability alone drives cell swelling and membrane lysis. We therefore demonstrate that a deeper understanding of these dynamic cell modifications and their consequences will shed light on the molecular and biophysical mechanisms driving different forms of cell death.

Abstract: Inflammasomes are multiprotein platforms that control caspase-1 activation, which process the inactive precursor forms of the inflammatory cytokines IL-1β and IL-18, leading to an inflammatory type of programmed cell death called pyroptosis. Studying inflammasome-driven processes, such as pyroptosis-induced cell swelling, under controlled conditions remains challenging because the signals that activate pyroptosis also stimulate other signaling pathways. We designed an optogenetic approach using a photo-oligomerizable inflammasome core adapter protein, apoptosis-associated speck-like containing a caspase recruitment domain (ASC), to temporally and quantitatively manipulate inflammasome activation. We demonstrated that inducing the light-sensitive oligomerization of ASC was sufficient to recapitulate the classical features of inflammasomes within minutes. This system showed that there were two phases of cell swelling during pyroptosis. This approach offers avenues for biophysical investigations into the intricate nature of cellular volume control and plasma membrane rupture during cell death.