Orthocaspases execute regulated cell death (RCD) in cyanobacteria, yet their post-stress activation and physiological relevance remain elusive. Here, we investigated this using heterologous expression of orthocaspases in Synechococcus elongatus PCC 7942. Orthocaspase-expressing (OE) strains, SynOC2GFP and SynOC6GFP, were generated using GFP-fused orthocaspases, AnaOC2 and AnaOC6. Catalytically inactive C/A variants were used to generate mutant-expressing (ME) controls, SynC140AGFP and SynC134AGFP. Under non-stress conditions, OE and ME strains accumulated orthocaspases without losing viability. In contrast, sulphur-limitation, heat, or cold stresses accelerated cell death in OE strains. Across all treatments orthocaspases activation was consistently initiated at ~15% DNA damage and peaked beyond ~30%, temporally coincided with recA expression. RecA and orthocaspases showed direct interactions, connecting stress-induced DNA damage and repair exhaustion to orthocaspases activation. Physiological relevance of the process was assessed by monitoring post-stress recovery in WT co-cultured with OE or ME strains. Population recovery after sulphur-limitation and cold stress occurred exclusively in WT + OE co-cultures. These regenerated populations were dominated by WT (85%-90%), indicating selective elimination of OE cells during stress. This suggested elimination of compromised cells by orthocaspases activity, limiting futile energy expenditure and propagation of mutation-prone genomes to progeny. Thus, orthocaspases-mediated cell death enhances inclusive fitness of cyanobacteria during abiotic stresses.
Bhattacharjee et al. (Wed,) studied this question.