Cell death upon exposure to darkness or <i>H.salinarum</i> cells triggers the release of glycerol by <i>D. salina</i>.

<p>(A) <i>D. salina</i> accumulates and utilizes glycerol as an osmoprotectant in hypersaline growth conditions. Accumulation of glycerol in <i>D. salina</i> cultures is correlated to increasing salinity in the growth medium peaking at 150 PSU (inset). (B) <i>D. salina</i> releases glycerol by cell death. Illustrated and merged phase contrast/fluorescence photomicrographs of a <i>D. salina</i> cell undergoing cell death. <i>D. salina</i> stores glycerol and other byproducts of photosynthesis inside secretory vesicles that are localized to the apical flagellar pole (top). The green color of the vesicles is due to quinacrine staining of glycerol and the red fluorescence corresponds to chloroplasts. The image show dramatic disruption of the cell membrane and complete loss of internal glycerol in a cell that has undergone cell death (bottom). (C) A shift of live light acclimated cultures (100–150 µmol-photon m⁻²sec⁻¹) to complete darkness (0 µmol-photon m⁻² sec⁻¹) results in release of glycerol by <i>D. salina</i>. The intracellular glycerol is measured by flow cytometry analysis of quinacrine stained vesicles <i>D. salina</i> cells. (D) Representative confocal laser micrograph of <i>D. salina</i> cells stained with FITC-Annexin V to highlight the externalization of PS (green fluorescence), and red corresponds to red chlorophyll autofluorescence. (E) Representative confocal laser micrographs of dead <i>D. salina</i> stained with FITC-Annexin V and SYTOX® blue highlighting PS completely externalized and the ejection of the nucleus indicating cell death.