Submitted to: Planta
Publication Type: Peer reviewed journal
Publication Acceptance Date: 11/10/2003
Publication Date: 3/10/2004
Citation: Tucker, D., Allen, D.J., Ort, D.R. 2004. Control of nitrate reductase by circadian and durral rhythm in tomato. Planta. 219:277-285. Interpretive Summary: The capacity of plant leaves to use photo synthetic energy to convert inorganic forms of nitrogen into compounds that can be incorporated in protein is a fundamental determinant of the nutritional quality of the harvest. The enzyme nitrate reductase plays a critical role in the overall of the metabolism of leaf nitrogen. The regulation of this enzyme is highly complex and responds to many environmental factors often making nitrate reductase the rate-controlling step in the production of amino acids, which are the building blocks of proteins. This work demonstrates that the regulation by the plant's 24 h biological clock (i.e., circadian clocks) of nitrate reductase gene activity has primary control of this enzyme's activity. The extensively studied control of nitrate reductase activity by protein phosphorylation changes is shown to play an ancillary role that is most important when coping with rapid changes in light level (e.g., intermittent cloud cover). These results are important to the effort by agricultural scientists to define how crop nitrogen metabolisms is controlled and limited in real environments.
Technical Abstract: Nitrate reductase (NR) (E.C. 184.108.40.206) is a key, regulatory step in the assimilation of nitrate into amino acids in plant leaves. NR activity is intricately controlled by multifarious regulatory mechanisms acting at different levels ranging from transcription to protein degradation. It is one of a few enzymes in plants known to have a robust circadian rhythm of enzyme activity. Although many aspects of NR regulation have been studied in depth, how these different types of control interact in a plant to deliver integrated control of activity in leaves under naturally varying light conditions over the course of the day has not been systematically investigated. This work documents that NR in young tomato leaves has an endogenous rhythm in mRNA and protein level, which in nearly all circumstances are in the phase with the rhythm in NR enzyme activity. Our data show that the diurnal control of NR activity in tomato leaves rests primarily with circadian regulation at the level of transcription. The accompanying oscillations in protein level are made possible by a short half-life of NR protein that is ~2.5 h under normal conditions. Ultimately the extensively studied NR post transcriptional regulation via phosphorylation and subsequent inhibitor protein binding has a physiologically vital but secondary, regulatory roles of rapidly deactivating NR in response changes in light intensity that cannot be anticipated by circadian timing. This deactivation appears to signal proteolytic degradation of NR that may be much more rapid than the 2.5 h half-life observed under circadian conditions. The post translational reactivation of NR appears to have a similarly limited physiological role in diurnal regulation of NR in tomato leaves.