CSIRO PUBLISHING www.publish.csiro.au/journals/fpb Functional Plant Biology, 2004, 31, 847–855 De novo protein synthesis in relation to ammonia and proline accumulation in water stressed white clover Tae-Hwan Kim A,E , Bok-Rye Lee A , Woo-Jin Jung B , Kil-Yong Kim C , Jean-Christophe Avice D and Alain Ourry D A Department of Animal Science, Institute of Agricultural Science and Technology, College of Agriculture and Life Science, Chonnam National University, Gwangju 500-757, Korea. B Glucosamine Saccharide Materials Laboratory (NRL), Institute of Agricultural Science and Technology, Chonnam National University, Gwangju 500-757, Korea. C Department of Biological and Environmental Chemistry, College of Agriculture and Life Science, APSRC, Chonnam National University, Gwangju 500-757, Korea. D UMR, INRA-UCBN, ´ Ecophysiologie v´ eg´ etale, Agronomie et Nutritions NCS, Institut de Biologie Fondamentale et Appliqu´ ee, Universit´ e de Caen, F-14 032 Caen Cedex, France. E Corresponding author; email: grassl@chonnam.ac.kr Abstract. The kinetics of protein incorporation from newly-absorbed nitrogen (N, de novo protein synthesis) was estimated by 15 N tracing in 18-week-old white clover plants (Trifolium repens L. cv. Regal) during 7 d of water-deficit treatment. The physiological relationship between kinetics and accumulation of proline and ammonia in response to the change in leaf-water parameters was also assessed. All leaf-water parameters measured decreased gradually under water deficit. Leaf and root dry mass was not significantly affected during the first 3d when decreases in leaf-water parameters were substantial. However, metabolic parameters such as total N, proline and ammonia were significantly affected within 1d of commencement of water-deficit treatment. Water-deficit treatment significantly increased the proline and NH 3 –NH 4 + concentrations in both leaves and roots. There was a marked reduction in the amount of N incorporated into the protein fraction from the newly absorbed N (NANP) in water-deficit stressed plants, particularly in leaf tissue. This reduction in NANP was strongly associated with an increased concentration of NH 3 –NH 4 + in roots (P≤0.05) and proline (P≤0.01) in leaves and roots. These results suggest that proline accumulation may be a sensitive biochemical indicator of plant water status and of the dynamics of de novo protein synthesis in response to stress severity. Keywords: ammonia, leaf-water parameters, 15 N-protein, proline, Trifolium repens, water-deficit stress. Introduction White clover has a low tolerance to drought because of its shallow root system. A prolonged drought, which can occur predominantly in spring in Korea, can kill a population, and high temperatures throughout the following summer are also unfavourable for growth. The consequence of poor sward clover content often leads to problems in agricultural practice. However, the mechanisms underlying the inhibition of plant growth by drought are not yet fully understood. During prolonged periods of drought, the decrease in water availability for transport-associated processes leads to Abbreviations used: NANP, the amounts of newly absorbed N in soluble protein; NCCs, nitrogen containing compounds; RSA, relative specific activities; RWC, relative water content;  π , leaf osmotic potential;  π 100 , leaf osmotic potential at full turgor;  w , leaf water potential;  soil , soil matric potential. changes in the concentrations of many metabolites, followed by disruptions to amino acid and carbohydrate metabolism. Several classes of compounds increase during drought, including compatible solutes such as the special amino acid proline, sugars, sugar-alcohols and glycinebetaine (Yancey et al. 1982; Girousse et al. 1996). Proposed roles for proline in stress tolerance include serving as osmotica, stabilising macromolecules and membranes and detoxifying tissues of excess N (Huang et al. 1994; Rabe 1999). Direct evidence supporting these roles was provided by some mutant and transgenic proline-over-producing plants, which show increased osmotolerance (Hong et al. 2000; Roosens et al. © CSIRO 2004 10.1071/FP04059 1445-4408/04/080847