Light-induced increase in free Mg2+ concentration in spinach chloroplasts: measurement of free Mg2+ by using a fluorescent probe and necessity of stromal alkalinization.
Free Mg(2+) in chloroplasts may contribute to the regulation of photosynthetic enzymes, but adequate methodology for the determination of free Mg(2+) concentration ([Mg(2+)]) in chloroplasts has been lacking. We measured internal chloroplast [Mg(2+)] by using a Mg-sensitive fluorescent indicator, mag-fura-2. In intact, dark-kept spinach chloroplasts, internal [Mg(2+)] was estimated to be 0.50 mM, and illumination caused an increase in [Mg(2+)] to 2.0mM in the stroma. The light-induced increase in [Mg(2+)] was inhibited by a blocker of driven electron transport and uncouplers. The K(+)-specific ionophore valinomycin inhibited the [Mg(2+)] increase in the absence of external K(+), and addition of KCl restored the [Mg(2+)] increase. NH(4)Cl, which induces stromal alkalinization, enhanced the [Mg(2+)] increase. A Ca(2+)-channel blocker, ruthenium red, inhibited the [Mg(2+)] increase, but LaCl(3) had no effect. These results indicate that stromal alkalinization is essential for light-induced increase in [Mg(2+)]. This system for measuring internal chloroplast [Mg(2+)] might provide a suitable system for assay of Mg(2+) transport activity of chloroplast membranes.
Ishijima
Sumio
S
Graduate School of Agriculture, The Prefecture University of Kyoto, Kyoto 606-8522, Japan. ishijima@kpu.ac.jp
Uchibori
Atsuhito
A
Takagi
Hiroki
H
Maki
Ryoji
R
Ohnishi
Masatake
M
D002118:Calcium / Q000378:metabolism; D002138:Calibration; D002736:Chloroplasts / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005456:Fluorescent Dyes / Q000494:pharmacology; D016257:Fura-2 / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D007811:Lanthanum / Q000494:pharmacology; D008027:Light; D008274:Magnesium / Q000494:pharmacology*; D011189:Potassium Chloride / Q000494:pharmacology; D012430:Ruthenium Red / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D018724:Spinacia oleracea / Q000378:metabolism*; D013997:Time Factors; D014634:Valinomycin / Q000494:pharmacology
D005456:Fluorescent Dyes; D007476:Ionophores; C028521:lanthanum chloride; D012430:Ruthenium Red; D014634:Valinomycin; D011189:Potassium Chloride; D007811:Lanthanum; D008274:Magnesium; D002118:Calcium; D016257:Fura-2