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

We investigated the possibility of Ca2+ signaling in cyanobacteria (blue-gr een algae) by measuring intracellular free Ca2+ levels ([Ca2+](i)) in a rec ombinant strain of the nitrogen fixing cyanobacterium Anabaena strain sp. P CC7120, which constitutively expresss the Ca2+-binding photoprotein apoaequ orin. The homeostasis of intracellular Ca2+ in response to increasing exter nal Ca has been studied in this strain. The resting level of free Ca2+ in A nabaena was found to be between 100 and 200 nM. Additions of increasing con centrations of external Ca2+ gave a transient burst of [Ca2+], followed by a very quick decline, reaching a plateau within seconds that brought the le vel of [Ca2+], back to the resting value. These results indicate that Anaba ena strain sp. PCC7120 is able to regulate its internal Ca2+ levels. We als o monitored Ca2+ transients in our recombinant strain in response to heat a nd cold shock. The cell's response to both stresses was dependent on the wa y they were induced. The use of inhibitors suggests that heat shock mobiliz es cytosolic Ca2+ from both intracellular and extracellular sources, while the Ca2+ source for cold shock signaling is mostly extracellular.