Programmed cell death is induced by hydrogen peroxide but not by excessive ionic stress of sodium chloride in the unicellular green alga Chlamydomonas reinhardtii

Eukaryotic microalgae serve as indicators of environmental change when exposed to severe seasonal fluctuations. Several environmental stress conditions are known to produce reactive oxygen species in cellular compartments, resulting in oxidative damage and apoptosis. The study of cell death in higher plants and animals has revealed the existence of an active ‘programmed cell death’ (PCD) process and similarities between such processes suggest an evolutionary origin. A study was undertaken to examine the morphological, biochemical and molecular responses of the unicellular green alga Chlamydomonas reinhardtii after exposure to oxidative (10 mM H₂O₂) and osmotic (200 mM NaCl and 360 mM sorbitol) stress. Concentrations of H₂O₂ (2–50 mM), NaCl and sorbitol (100–800 mM) were negatively correlated with growth. Biochemical analyses showed an increase in intracellular H₂O₂ production (2.2-fold with H₂O₂ and ~1.2–1.4-fold with NaCl and sorbitol) and activities of some antioxidant enzymes [super oxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX)]. Alteration of mitochondrial membrane potential (MMP) was observed upon treatment with H₂O₂ and NaCl, but not with sorbitol, indicating that the ionic stress component of NaCl altered the MMP. In addition, H₂O₂ led to the activation of a caspase-3-like protein, increase in the cleavage of a poly(ADP) ribose polymerase-1 (PARP-1)-like enzyme and formation of DNA nicks and laddering. With NaCl and sorbitol, no caspase activation, nor oligonucleosomal DNA laddering was observed, indicating non-apoptotic death. However, genomic DNA of NaCl (800 mM)-stressed cells, but not those of sorbitol-treated cells showed complete shearing. We conclude that the ionic rather than the osmotic component of NaCl leads to necrosis. These results unequivocally suggest that the vegetative cells of C. reinhardtii respond differentially to various stress agents, leading to different death types in the same organism. Moreover, unlike most other organisms, when exposed to NaCl this alga does not undergo PCD.