Coaction of light, nitrate and a plastidic factor in controlling nitrite-reductase gene expression in tobacco

Abstract

Nitrite reductase (NIR; EC 1.7.7.1) - a key enzyme of nitrate reduction - is known to be induced by nitrate and light. In the present study with tobacco (Nicotiana tabacum L.) seedlings the dependency of NIR gene expression on nitrate, light and a plastidic factor was investigated to establish the nature of the coaction between these controlling factors. A cDNA clone coding for tobacco plastidic NIR was available as a probe. The major results were as follows: (i) The light effect on the appearance of NIR occurred predominantly through phytochrome. However, a specific blue-light effect was also involved. (ii) There was no effect of light on NIR appearance in the absence of nitrate while light exerted a strong effect when nitrate was provided. (iii) Anion-exchange chromatography revealed only a single form of NIR. While experiments involving plastid photooxidation indicated that this NIR is plastidic, a small residual level could not be eliminated by photooxidation. (iv) Northern blot analysis of NIR-transcript levels indicated that a low transcript level existed in the absence of nitrate and light; however, this level appeared to be increased slightly by light (in the absence of nitrate) and by nitrate (in the absence of light). A high transcript level was detected only when light as well as nitrate were provided. A low level was found when the plastids were damaged by photooxidation. It is concluded that plastidic NIR gene expression in tobacco requires positive control by a plastidic factor. Moreover, a synergistic action of phytochrome and nitrate is required to bring about a high transcript level. As found previously with mustard and spinach seedlings, there is no quantitative relationship between the transcript level and the rate of enzyme synthesis.