Heat-stress-induced programmed cell death in Heterosigma akashiwo (Raphidophyceae)

Programmed cell death (PCD) was previously thought to only occur in metazoans. Recent studies have revealed PCD in yeast, prokaryotes, and some species of phytoplankton. However, the need for PCD assays with reliable controls currently limits the study of PCD in natural environments. We examined heat-stressed cultures of Heterosigma akashiwo (Y. Hada) Y. Hada ex Y. Hara and M. Chihara for DNA fragmentation (using the terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay), externalization of phosphatidylserine residues (using Annexin-V) and caspase-like activity (using CaspACE and Caspase-Glo 8), which are all diagnostic of cells undergoing PCD but not necrosis. Additionally, we treated cultures with the broad-spectrum caspase inhibitor z-VAD-FMK to rescue cells otherwise destined for death. Our results demonstrate the range of inducible responses of H. akashiwo to heat-stress: recovery from 35 °C heat-stress, PCD-induction due to 37 °C and 40 °C heat-stress, and necrotic death or cyst formation due to 50 °C heat-stress. The identification of a reliable inducer and markers for PCD provides key tools for further studies into the physiology and ecology of this harmful bloom-former.

► Programmed cell death occurs in certain phytoplankton species. ► A range of responses was observed in Heterosigma akashiwo due to heat stress. ► Recovery, PCD and necrosis occurred after mild, moderate and severe stress, respectively. ► Identification of reliable markers and detection methods will now permit further study.