Physiological characterization of a plastidic signal required for nitrate-induced appearance of nitrate and nitrite reductases

Abstract

We compared the response of NO 3 (-) -induced nitrate-reductase (NR) and nitrite-reductase (NIR) levels in virtually carotenoid-free far-red-light-grown mustard (Sinapis alba L.) cotyledons following a photooxidative treatment of the plastids. The cytosolic localization of NR and the plastidic localization of NIR were confirmed with this approach. Emphasis was on a plastidic factor previously postulated to be involved obligatorily in the transcriptional control of nuclear genes coding for proteins destined for the chloroplast. Photooxidative damage of the plastid would be to destroy the ability of the organelle to send off this signal. Dependency of NIR and NR induction by NO 3 (-) on the plastidic factor is described in detail, and it is concluded that requirement for the plastidic factor is relatively high in the case of NR while factor requirement to allow induction is low in the case of NIR. The data indicate that in the case of NIR the photooxidative damage done to the plastid also affects accumulation of the enzyme directly. Since this effect is absent in the case of cytosolic NR, induction of NR is a particularly suitable system for further molecular studies of the plastidic factor and its mode of action.

The Role of Plastids in the Expression of Nuclear Genes for Thylakoid Proteins Studied with Chimeric [beta]-Glucuronidase Gene Fusions

“…The activities of the stromal NADP+-dependent glyceraldehyde-3-phosphate dehydrogenase and ribulose-1,5-bisphosphate carboxylase were at least 20-fold reduced (1.4 and 4.6%, respectively, of the corresponding water controls) and that of NiR was approximately 10-fold lower (9.4% of the water control), but the activities of the cytoplasmic NAD+-dependent glyceraldehyde-3-phosphate dehydrogenase and malate dehydrogenase were slightly increased (1 13 and 126%, respectively). The only exception is the cytoplasmic NR activity, which is not detectable in photobleached seedlings (less than 1 %), consistent with previous reports from other species (Borner et al, 1986;Rajasekhar and Oelmiiller, 1987;Oelmiiller et al, 1988). Quantitation of dot blot signals from RNA hybridized to probes specific for plastid genes demonstrate that the steady-state transcript levels of the rDNA, psbA, psbE, petA, and atpA genes are approximately 20-fold lower compared to the water controls and comparable to the levels found in tobacco seeds, whereas etiolated seedlings contain almost identical or slightly reduced Northem analyses with cDNAs for six thylakoid membrane proteins demonstrate that the transcript levels of three polypeptides associated with Chls (CAB I, CAB 11, and the CP 24 apoprotein; genes CAB I, CAB 11, CP 24, respectively) are below or at the limits of detectability in herbicide-treated seedlings, whereas those for the Rieske Fe/S polypeptide (PETC), as well as for the 33-kD (PSBO) and 23-kD (PSBP) polypeptides, were only moderately reduced ( Fig.…”

“…An additional complexity derives from the observation that transcript levels of nuclear genes coding for cytoplasmic enzymes with functions related to plastids, such as NR, also fail to be expressed in tissues with impaired plastids (Bomer et al, 1986;Oelmiiller et al, 1988;Oelmiiller and Briggs, 1989). However, this might be caused by a completely different regulatory network.…”

Photooxidative Destruction of Chloroplasts and Its Effect on Nuclear Gene Expression and Extraplastidic Enzyme Levels *

“…1973;Frosch et al, 1979;Batschauer et al, 1986;Oelmiiller et al, 1986bOelmiiller et al, , 1988Schuster et al, 1988b). However, as soon as all Chl is photooxidized, no further photodamage occurs even if the seedlings are maintained in strong light (Frosch et al, 1979;Oelmiiller et al, 1988).…”

On The Regulation of Spinach Nitrate Reductase

“…NR3 can be regarded as a key enzyme in this process. In spite of a contradicting report (14), most evidence supports the view that nitrate reductase is located in the cytosolic compartment (2,21,32). As sources of redox equivalents for the generation of NADH required for nitrate reduction, the oxidation of glyceraldehyde-3-phosphate via the cytosolic NAD-GAPDH (15) and/or the oxidation of malate catalyzed by the cytosolic MDH (17) have been discussed.…”