Nitric oxide production mediated by nitrate reductase in the green alga Chlamydomonas reinhardtii: an alternative NO production pathway in photosynthetic organisms.
Biological activity of nitric oxide (NO) production was investigated in the unicellular green alga Chlamydomonas reinhardtii. An NO specific electrode detected a rapid increase in signal when nitrite (NO(2)(-)) was added into a suspension of C. reinhardtii intact cells in the dark. The addition of KCN or the NO quencher bovine hemoglobin completely abolished the signal, verifying that the nitrite-dependent increase in signal is due to enzymatic NO production. L-arginine, the substrate for NO synthase, did not induce detectable NO production and the NOS inhibitor N(omega)-nitro-L-arginine showed no inhibitory effect on the nitrite-dependent production of NO. Illuminating cells showed a significant suppressive effect on NO production. When the photosynthetic electron transport inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea was present in the suspension, C. reinhardtii cells produced NO after the addition of nitrite even under illumination. Kinetic and microscopic observations, using the intracellular fluorescent NO probe 4,5-diaminofluorescein-2 diacetate, both demonstrated that NO was produced within the cells in response to the addition of nitrite. The Chlamydomonas mutant cc-2929, which lacks nitrate reductase (NR) activity, did not display any of the responses observed in the wild-type cells. The results presented here provide direct in vivo evidence to confirm that NR is involved in the nitrite-dependent NO production in the green alga.
Sakihama
Yasuko
Y
Laboratory of Cell and Functional Biology, Faculty of Science, University of the Ryukyus, Nishihara, Okinawa, 903-0213, Japan.
Nakamura
Soichi
S
Yamasaki
Hideo
H
D000818:Animals; D016825:Chlamydomonas reinhardtii / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D004237:Diuron / Q000494:pharmacology; D019793:Fluorescein / Q000494:pharmacology; D008958:Models, Molecular; D009154:Mutation; D050901:Nitrate Reductase; D009565:Nitrate Reductases / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D010101:Oxygen Consumption; D010788:Photosynthesis / Q000502:physiology; D011190:Potassium Cyanide / Q000494:pharmacology; D012977:Sodium Nitrite / Q000494:pharmacology
C414689:4,5-diaminofluorescein diacetate; D009569:Nitric Oxide; D004237:Diuron; D019001:Nitric Oxide Synthase; D009565:Nitrate Reductases; D050901:Nitrate Reductase; D012977:Sodium Nitrite; D011190:Potassium Cyanide; D019793:Fluorescein