Finally, we triggered mild optoDronc activation in clones and found that their death rate 1.5-5hr later was 3.5-fold that of their WT neighbours, indicating that sublethal caspase activity can bias clone elimination in vivo.

Interestingly, when we block prior caspase with RNAi and trigger optoDronc, there is still a bias towards increased death at the midline and anterior side of the tissue, indicating other unknown spatially patterned factors modulating sensitivity to caspase independently of past activation.

To test if such priming occurs, we used optoDronc to subject one lateral domain of the tissue to two pulses of mild caspase activation separated by a number of hours (blue box), and measured the number of deaths against the control contralateral domain only subjected to the latter pulse (red box).

To test the physiological relevance of this cell-to-cell heterogeneity in caspase threshold, we used optoDronc, an optogenetic caspase, to trigger mild caspase activation everywhere in the notum and track where cells were most and least likely to die.

By detecting the point at which individual tracked cells irreversibly commit to apoptotic extrusion, we searched for a shared threshold of cumulative (GC3Ai) or instantaneous (δGC3Ai) caspase activity that defines this point of commitment.

The bifurcation between survival and death was generally assumed to be driven by a threshold of caspase activity, but this was never tested quantitatively in vivo. We tracked over 15,000 cells expressing a caspase activity sensor in a developing epithelium to probe the existence of such a threshold.

The effector caspases drive cell death, but their activation can often be survived, so how do cells make this decision? Our new preprint from @levayerr.bsky.social shows that instantaneous caspase activity is important, but past activation has a key role to play!

www.biorxiv.org/content/10.1...