Copper-induced calcium release from ER involves the activation of ryanodine-sensitive and IP3-sensitive channels in Ulva compressa

The marine alga Ulva compressa (Chlorophyta) showed a triphasic release of intracellular calcium with maximal levels at 2, 3 and 12 h and a biphasic accumulation of intracellular hydrogen peroxide with peaks at 3 and 12 h when cultivated with copper excess. Intracellular hydrogen peroxide originated exclusively in organelles. In this work, we analyzed the intracellular origin of calcium release and the type of calcium channels activated in response to copper excess. U. compressa was treated with thapsigargin, an inhibitor of endoplasmic reticulum (ER) calcium ATPase, ryanodine, an inhibitor of ryanodine-sensitive channels and xestospongin C, an inhibitor of inositol 1, 4, 5-triphosphate (IP₃)-sensitive channels. Thapsigargin induced the depletion of calcium stored in ER at 75 min and completely inhibited calcium release at 2, 3 and 12 h of copper exposure indicating that calcium release originated in ER. In addition, ryanodine and xestospogin C inhibited calcium release at 2 and 3 h of copper exposure whereas the peak at 12 h was only inhibited by ryanodine. Thus, copper induced the activation of ryanodine-sensitive and IP₃-sensitive calcium channels in ER of U. compressa.Key words: calcium release, endoplasmic reticulum, calcium channels, marine alga, Ulva compressaPlants showed common responses to biotic and abiotic stresses, mainly the accumulation of reactive oxygen species (ROS), in particular hydrogen peroxide, and the release of intracellular calcium.^1,2 Regarding abiotic stress, it has been shown that ozone triggers a NADPH oxidase-dependent biphasic oxidative burst in Arabidopsis thaliana that activates antioxidant and defense enzymes.^3,4 In addition, cadmium induced a NADPH oxidase-dependent monophasic accumulation of extracellular hydrogen peroxide in tobacco cells.⁵ On the other hand, ozone as well as absicic acid treatment, dessication, cold, heat, salinity, UV light and anoxia induce intracellular calcium release and the activation of antioxidant enzymes.^6–8 Regarding abiotic stress in algae, copper induced a monophasic increase of intracellular hydrogen peroxide at 2 h of copper exposure in the brown seaweeds Lessonia nigrecsens and Scytosiphon lomentaria.⁹ On the other hand, strontium induced calcium release in the green microalga Eremosphaera viridis as did osmotic stress in the zygote of the brown macroalga Fucus serratus.^10,11 U. compressa is a cosmopolitan marine macroalga (Chlorophyta) growing in copper-impacted coastal areas in northern Chile.¹² U. compressa cultivated in seawater with copper excess (10 µM) showed co-occuring increases of intracellular calcium and hydrogen peroxide.¹³ Copper induced a triphasic release of calcium with maximal levels at 2, 3 and 12 h and a biphasic production of hydrogen peroxide with peaks at 3 and 12 h. Interestingly, the production of hydrogen peroxide occurred exclusively in organelles, i.e., mitochondria and chloroplasts. In addition, calcium and hydrogen peroxide act as signals in the differential activation of antioxidant and defense enzymes.¹³ In this work, we analyzed the intracellular origin of copper-induced calcium release and the type of calcium channels activated in response to copper excess in U. compressa.