Use of recombinant aequorin to study calcium homeostasis and monitor calcium transients in response to heat and cold shock in cyanobacteria

Abstract

We investigated the possibility of Ca2+ signaling in cyanobacteria (blue-green algae) by measuring intracellular free Ca2+ levels ([Ca2+](i)) in a recombinant strain of the nitrogen fixing cyanobacterium Anabaena strain sp. PCC7120, which constitutively expresses the Ca2+-binding photoprotein apoaequorin. The homeostasis of intracellular Ca2+ in response to increasing external Ca2+ has been studied in this strain. The resting level of free Ca2+ in Anabaena was found to be between 100 and 200 nM. Additions of increasing concentrations of external Ca2+ gave a transient burst of [Ca2+](i) followed by a very quick decline, reaching a plateau within seconds that brought the level of [Ca2+](i) back to the resting value. These results indicate that Anabaena strain sp. PCC7120 is able to regulate its internal Ca2+ levels. We also monitored Ca2+ transients in our recombinant strain in response to heat and cold shock. The cell's response to both stresses was dependent on the way they were induced. The use of inhibitors suggests that heat shock mobilizes cytosolic Ca2+ from both intracellular and extracellular sources, while the Ca2+ source for cold shock signaling is mostly extracellular.