Organic acids play a pivotal role in improving plant response to long‐term drought stress. External application of organic acids has been reported to improve drought resistance in several species. However, whether organic acids have similar effects in tobacco remains unknown. A screening study of the protective function of organic acids in tobacco and understanding the underlying molecular mechanism would be useful in developing a strategy for drought tolerance. Several physiological and molecular adaptations to drought including abscisic acid, stomatal closure, reactive oxygen species homeostasis, amino acid accumulation, and drought‐responsive gene expression were observed by exogenous citric acid in tobacco plants. Exogenous application of 50 mm citric acid to tobacco plants resulted in higher chlorophyll content, net photosynthesis, relative water content, abscisic acid content and lower stomatal conductance, transpiration and water loss under drought conditions. Moreover, reactive oxygen species homeostasis was better maintained through increasing activity of antioxidant enzymes and decreasing hydrogen peroxide content after citric acid pretreatment under drought. Amino acids involved in the TCA cycle accumulated after external application of citric acid under drought stress. Furthermore, several drought stress‐responsive genes also dramatically changed after application of citric acid. These data support the idea that external application of citric acid enhances drought resistance by affecting physiological and molecular regulation in tobacco. This study provides clear insights into mechanistic details of regulation of amino acid and stress‐responsive gene expression by citric acid in tobacco in response to drought, which is promising for minimizing growth inhibition in agricultural fields.

Biosynthesis of galactolipids in spinach chloroplasts has been reported (2,5). In these studies uridine diphosphate galactose was used as a galactosyl donor to some endogenous acceptor (diglyceride) (Branson Instruments Inc.). Chloroplasts were isolated from the broken cells by the procedure described by Carell and Kahn (Flb fraction) (1). The purity of the chloroplasts was checked under a microscope. Enzymatic synthesis of galactolipids was carried out by the method described previously (2). A typical reaction mixture contained, in 1 ml: uridine diphosphate galactose ("C) (0.03 lac giving 24,000 cpm with specific radioactivity of 25 Ac ,Mmole) in 0.1 ml; chloroplasts or broken cells (dark-grown cells) equivalent to about 5 mg of proteins in 0.5 ml; 0.4 ml of 0.1 M tris buffer, pH 7.4. At the end of a 1-hr reaction at 37 C, the reaction was stopped by adding 4 ml of chloroform-methanol (2:1 v/v) and heating at 55 C for 5 min. The lipid products were extracted once more with 2 ml of the chloroform-methanol solution. The lipid extract was washed four times with 2.5 ml of 0.9%c aqueous sodium chloride solution to remove residual radioactive uridine diphosphate galactose. A 1-ml aliquot of the lipid extract was transferred to a planchet, dried, and counted. The rest of the lipid extract was concentrated and separated on a column packed with silicic acid (Unisil, Clarkson Co., Williamsport, Penn.) into monogalactosyl diglyceride and digalactosyl diglyceride by eluting the column with 100 ml of a solution of 5% diethyl ether in methanol and equal volume of 20% diethyl ether in methanol, respectively. Each eluate was concentrated and counted on a planchet by means of a gas flow counter (Nuclear-Chicago).