Involvement of calcium mobilization from caffeine-sensitive stores in mechanically induced cell cycle arrest in the dinoflagellate Crypthecodinium cohnii.

摘要

Mechanical loads can profoundly alter cell growth and cell proliferation. The dinoflagellates are especially sensitive to mechanical stimulation. Many species will be arrested in cell cycle in response to turbulence or shear stress. We demonstrate here that mechanical shaking and caffeine, the ryanodine-receptor agonist, induced an elevation of cytosolic calcium in the dinoflagellate Crypthecodinium cohnii. Dantrolene, a ryanodine-receptor antagonist, dose-dependently inhibited both shaking-induced and caffeine-induced calcium release. Similar to the effect of mechanical shaking, caffeine alone dose-dependently and reversibly induced cell cycle arrest in dinoflagellates. Prolonged shaking substantially abolished the magnitude of caffeine-induced calcium release and vice-versa, suggesting that both agents released calcium from similar stores through ryanodine receptors. Fluorescence-conjugated ryanodine gave positive labeling, which could be blocked by ryanodine, in the cortice of C. cohnii cells. In addition, caffeine or shaking mobilized intracellular chlortetracycline (CTC)-positive membrane-bound calcium, which could be similarly depleted by t-BuBHQ, a SERCA pump inhibitor. Prior treatment with shaking or caffeine also inhibited the ability of the other agent in mobilizing CTC-positive calcium. CTC-positive microsomal fractions could also be induced to release calcium by caffeine and cADPR, the ryanodinee receptor modulator. t-BuBHQ, but not calcium ionophores, induced cell cycle arrest, and the calcium chelator BAPTA-AM was unable to rescue caffeine-induced cell cycle arrest. These data culminate to suggest that mobilization or depletion of caffeine-sensitive calcium stores, but not calcium elevation per se, is involved in the induction of cell cycle arrest by mechanical stimulation. The present study establishes the role of caffeine-sensitive calcium stores in the regulation of cell cycle progression.