title: Loading of quin2 into the oat protoplast and measurement of cytosolic calcium ion concentration changes by phytochrome action.
abstract: The loading of quin2 into oat protoplasts was carried out in an incubation medium (0.6 M sorbitol, 1 mM CaCl2, 5 mM Mes, 5 mM Tris, 0.05% BSA, 1 mM KCl, 1 mM MgSO4 (pH 6.8)), in which we found the best viability of the protoplast and the highest membrane permeability of quin2/AM, compared with the results obtained from any other incubation medium we had tried to use. 50 microns of quin2/AM was added in the suspension medium containing 5 x 10(5)/ml of oat protoplasts, and incubation at 4 degrees C was performed for 24 h. From atomic absorption data, we confirmed that quin2 loading was 1.78 mmol per liter of cells. Red-light (660 nm) irradiation for 5 min caused an increase of the cytosolic Ca2+ concentration from 30 to 193 nM. On the other hand, a subsequent irradiation with far-red light (730 nm) for 5 min decreased it by about 48 nM. Even when the extracellular Ca2+ was completely chelated with 1 mM EDTA, red light increased the cytosolic Ca2+ concentration by about 51 nM and far-red light decreased it to 3 nM. These results imply that the Pfr form of phytochrome functions not only in the process of influx of Ca2+, but also in the mobilization process of Ca2+ from the intracellular Ca2+ pools. The fact that the Pr form of phytochrome lowers the cytosolic Ca2+ concentration is also presented in this report.
authors.affiliation: Department of Biochemistry, Chungbuk National University, Cheongju, Korea.
authors.forename: Q
authors.initials: Q
authors.lastname: Chae
authors.forename: H J
authors.initials: HJ
authors.lastname: Park
authors.forename: S D
authors.initials: SD
authors.lastname: Hong
mesh_terms: D000634:Aminoquinolines / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D002523:Edible Grain; D005456:Fluorescent Dyes; D010834:Phytochrome / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism*; D011523:Protoplasts / Q000187:drug effects / Q000378:metabolism*
chemical_list: D000634:Aminoquinolines; D005456:Fluorescent Dyes; D010940:Plant Proteins; D010834:Phytochrome; C035738:Quin2-acetoxymethyl ester; D002118:Calcium