ABSTRACT
An antisense nitrite reductase (NiR, EC 1.7.7.1) tobacco ( Nicotiana tabacum L.) transformant (clone 271) was used to gain insight into a possible correlation between nitrate reductase (NR, EC 1.6.6.1)-dependent nitrite accumulation and nitric oxide (NO(.)) production, and to assess the regulation of signal transduction in response to stress conditions. Nitrite concentrations of clone 271 leaves were 10-fold, and NO(.) emission rates were 100-fold higher than in wild type leaves. Increased protein tyrosine nitration in clone 271 suggests that high NO(.) production resulted in increased peroxynitrite (ONOO(-)) formation. Tyrosine nitration was also observed in vitro by adding peroxynitrite to leaf extracts. As in mammalian cells, NO(.) and derivatives also increased synthesis of proteins like 14-3-3 and cyclophilins, which are both involved in regulation of activity and stability of enzymes.
Subject(s)
Nicotiana/genetics , Nitric Oxide/biosynthesis , Nitrite Reductases/metabolism , Nitrites/metabolism , Signal Transduction/physiology , 14-3-3 Proteins , Antisense Elements (Genetics)/genetics , Carbon Dioxide/metabolism , Cyclophilins/biosynthesis , Ferredoxin-Nitrite Reductase , Light , Nitrate Reductase (NADH) , Nitrate Reductases/metabolism , Nitrite Reductases/genetics , Peroxynitrous Acid/metabolism , Peroxynitrous Acid/pharmacology , Plants, Genetically Modified , Signal Transduction/genetics , Nicotiana/metabolism , Tyrosine/drug effects , Tyrosine/metabolism , Tyrosine 3-Monooxygenase/biosynthesisABSTRACT
Blue light controls the development of sporangiophores in the zygomycete Phycomyces blakesleeanus Burgeff. Light represses the production of microsporangiophores and enhances the development of macrosporangiophores. Inhibition of the biosynthesis of tetrahydrobiopterin, a cofactor of NO synthase, inhibits this photomorphogenesis. Light induces production of citrulline from arginine in the mycelium and in sporangiophores. The citrulline-forming activity is dependent on NADPH, independent of calcium, and inhibited by NO synthase inhibitors. It is reduced in tetrahydrobiopterin-depleted mycelium. Light induces emission of NO from the developing fungus in the same order of magnitude as citrulline formation from arginine. The NO donor sodium nitroprusside can replace the light effect on sporangiophore development, and inhibitors of NO synthase repress it. We suggest that a fungal NO synthase is involved in sporangiophore development and propose its participation in light signaling.
Subject(s)
Biopterins/analogs & derivatives , Nitric Oxide Synthase/physiology , Phycomyces/enzymology , Biopterins/metabolism , Citrulline/metabolism , Culture Media , Enzyme Inhibitors/pharmacology , Light , Nitric Oxide/metabolism , Nitric Oxide Synthase/antagonists & inhibitors , Phycomyces/growth & development , Phycomyces/radiation effects , Signal Transduction , Spores, Fungal/physiologyABSTRACT
Purified plasma membranes (PMs) of tobacco (Nicotiana tabacum L. cv. Samsun) roots exhibited a nitrite-reducing enzyme activity that resulted in nitric oxide (NO) formation. This enzyme activity was not detected in soluble protein fractions or in PM vesicles of leaves. At the pH optimum of pH 6.0, nitrite was reduced to NO with reduced cytochrome c as electron donor at a rate comparable to the nitrate-reducing activity of root-specific succinate-dependent PM-bound nitrate reductase (PM-NR). The hitherto unknown PM-bound nitrite: NO-reductase (NI-NOR) was insensitive to cyanide and anti-NR IgG and thereby proven to be different from PM-NR. Furthermore, PM-NR and NI-NOR were separated by gel-filtration chromatography and apparent molecular masses of 310 kDa for NI-NOR and 200 kDa for PM-NR were estimated. The PM-associated NI-NOR may reduce the apoplastic nitrite produced by PM-NR in vivo and may play a role in nitrate signalling via NO formation.
