One‐fifth of irrigated agriculture is adversely affected by soil salinity. Hence, developing salt‐tolerant crops is essential for sustaining food production. Progress in breeding for salt‐tolerant crops has been hampered by the lack of understanding of the molecular basis of salt tolerance and lack of availability of genes that confer salt tolerance. Genetic evidence suggests that perception of salt stress leads to a cytosolic calcium‐signal that activates the calcium sensor protein SOS3. SOS3 binds to and activates a ser/thr protein kinase SOS2. The activated SOS2 kinase regulates activities of SOS1, a plasma membrane Na+/H+ antiporter, and NHX1, a tonoplast Na+/H+ antiporter. This results in Na+ efflux and vacuolar compartmentation. A putative osmosensory histidine kinase (AtHK1)‐MAPK cascade probably regulates osmotic homeostasis and ROS scavenging. Osmotic stress and ABA (abscisic acid)‐mediated regulation of LEA (late‐embryogenesis‐abundant)‐type proteins also play important roles in plant salt tolerance. Genetic engineering of ion transporters and their regulators, and of the CBF (C‐repeat‐binding factor) regulons, holds promise for future development of salt‐tolerant crops.

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Susceptibility to chilling injury prevents the cultivation of many important crops and limits the extended storage of horticultural commodities. Although freezing tolerance is acquired through cold-induced gene expression changes mediated in part by the CBF family of transcriptional activators, whether plant chilling resistance or sensitivity involves the CBF genes is not known. We report here that an Arabidopsis thaliana mutant impaired in the coldregulated expression of CBF genes and their downstream target genes is sensitive to chilling stress. Expression of CBF3 under a strong constitutive promoter restores chilling resistance to the mutant plants. The mutated gene was cloned and found to encode a nuclear localized RNA helicase. Our results identify a regulator of CBF genes, and demonstrate the importance of gene regulation and the CBF transcriptional activators in plant chilling resistance. L ow temperature is an important environmental factor that greatly influences the growth, development, survival, and distribution of plants (1). Most plants from temperate regions can cold-acclimate, i.e., they show increased tolerance to freezing temperatures after an exposure to low, nonfreezing temperatures (2). At least part of the basis of cold acclimation is that exposure to low temperatures induces the expression of many plant genes (2, 3). Some of the cold-induced gene products such as COR15A have been shown to mitigate membrane damage caused by freezing stress (4, 5).The dehydration-responsive element (DRE)͞C-repeat (CRT) cis-element is present in the promoters of many of the coldresponsive genes such as RD29A (also known as COR78 or LTI78) and COR15A (6-8). In Arabidopsis thaliana, the CBF family of transcriptional activators, also known as DREB1s, bind to the DRE͞CRT element and activate the expression of COR͞RD genes (8, 9). Cold induces rapid and transient expression of CBF1, CBF2, and CBF3, which in turn activate the downstream COR genes (10). Ectopic expression of CBF1 or CBF3 in transgenic Arabidopsis plants leads to constitutive expression of COR genes and enhanced freezing tolerance without cold-acclimation treatment (9, 11). These studies demonstrate a critical role of the CBF regulon in the acquisition of freezing tolerance. An important challenge ahead is to identify regulators of the CBF genes.Recently, the Arabidopsis HOS1 protein was shown to be a negative regulator of CBF genes (12). CBFs and their downstream COR genes show enhanced cold induction in hos1 mutant plants (12, 13). The Arabidopsis esk1 mutants are constitutively freezing-tolerant, but are not affected in the expression of genes with the DRE͞CRT cis-element, suggesting that ESK1 may be involved in a CBF-independent cold-response pathway (14). The sfr6 mutation reduces cold and abscisic acid (ABA) induction of the CRT͞DRE genes but has no effect on the expression of CBF genes (15). Therefore, no positive regulator of CBFs has been identified to date.Many important crops and fruits that originated from the tropics or subtropics, such a...

Previous workl-5 has shown the existence of a high-energy condition of isolated chloroplasts, caused by illumination in the absence of phosphate or ADP. This state is inferred from the ability to form ATP in the postillumination darkness. Accompanying the condition is an apparent uptake of hydrogen ions6z I together with an excretion of Mg++ or other cations.8 A feasible interpretation, stemming from the "chemi-osmotic" hypothesis for the mechanism of phosphorylation in double membrane containing organelles,9' 10 is that illumination causes the uptake of hydrogen ions into the inner space of the grana disk double membranes. The resulting inequality in hydrogen ion electrochemical activity across the membrane is postulated to be, in itself, the high-energy condition, able to drive the formation of ATP. If this interpretation is correct, then the same high-energy condition should be formed artificially, entirely in the dark and without electron transport, by loading the inner space of the grana disk membranes with protons. If this could be accomplished by placing chloroplasts in an acid medium, when first returned to pH 8 they might be expected to make some ATP due to the pH gradient across the membranes. The operational formation of ATP in this manner, entirely in the dark, was discovered and noted briefly earlier.4 The present paper represents an extension and further exploration of this phenomenon of "acid-bath" dark phosphorylation by chloroplasts. Materiats and Methods.-Chloroplasts were prepared from market spinach as described pre

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Betaine is an important osmoprotectant, synthesized by many plants in response to abiotic stresses. Almost all known biosynthetic pathways of betaine are two-step oxidations of choline. Recently, a biosynthetic pathway of betaine from glycine, catalyzed by two N-methyltransferase enzymes, was found. Here, the potential role of N-methyltransferase genes for betaine synthesis was examined in a freshwater cyanobacterium, Synechococcus sp. PCC 7942, and in Arabidopsis plants. It was found that the coexpression of N-methyltransferase genes in Synechococcus caused accumulation of a significant amount of betaine and conferred salt tolerance to a freshwater cyanobacterium sufficient for it to become capable of growth in seawater. Arabidopsis plants expressing N-methyltransferase genes also accumulated betaine to a high level in roots, stems, leaves, and flowers and improved seed yield under stress conditions. Betaine levels were higher than those produced by choline-oxidizing enzymes. These results demonstrate the usefulness of glycine N-methyltransferase genes for the improvement of abiotic stress tolerance in crop plants.cyanobacteria ͉ methyltransferase ͉ osmoprotectant ͉ stress resistance T oday, Ϸ20% of the world's cultivated land and nearly half of all irrigated lands are affected by high salinity (1). High salinity causes ion imbalance and hyperosmotic stress in plants. Organisms that thrive in hypersaline environments possess specific mechanisms for the adjustment of their internal osmotic status. One such mechanism is the ability to accumulate low-molecular-weight organic-compatible solutes such as sugars, some amino acids, and quaternary ammonium compounds (2-4). Glycine betaine (N,N,Ntrimethylglycine, hereafter betaine) is a major osmolyte (2-4). Another mechanism for adaptation to high salinity is the exclusion of the Na ϩ ion from sodium-sensitive sites (5). Genetic engineering techniques have been applied to improve the salt tolerance of plants (6-13). Considerable success has been demonstrated by manipulating the Na ϩ ͞H ϩ antiporter genes (6-8). By contrast, the genetic engineering of betaine synthesis has been hampered by low accumulation levels of betaine (9-13). Most known biosynthetic pathways of betaine include a two-step oxidation of choline: choline 3 betaine aldehyde 3 betaine. The first step is catalyzed by choline monooxygenase (CMO) in plants (14), choline dehydrogenase (CDH) in animals and bacteria (15,16), and choline oxidase in some bacteria (11,17). The second step is catalyzed by NAD ϩ -dependent betaine aldehyde dehydrogenase in all organisms (15,18,19), although in some bacteria, CDH and choline oxidase also catalyze the second step (15-17). Hitherto, all attempts at betaine synthesis have been carried out by using choline-oxidizing enzymes (9-13). The supply and transport of betaine precursors such as choline, ethanolamine, and serine to plastids may be of importance, because these precursors have been suggested to be limiting (12,13).Recently, we showed that a halotolerant cyanobacter...

Transgenic tobacco plants expressing the ascorbate oxidase (AAO) gene in sense and antisense orientations, and an Arabidopsis mutant in which the T-DNA was inserted into a putative AAO gene, were used to examine the potential roles of AAO for salt-stress tolerance in plants. AAO activities in the transgenic tobacco plants expressing the gene in sense and antisense orientations were, respectively, about 16-fold and 0.2-fold of those in the wild type. Under normal growth conditions, no significant differences in phenotypes were observed, except for a delay in flowering time in the antisense plants. However, at high salinity, the percentage germination, photosynthetic activity, and seed yields were higher in antisense plants, with progressively lower levels in the wild type and the sense plants. The redox state of apoplastic ascorbate in sense plants was very low even under normal growth conditions. Upon salt stress, the redox state of symplastic and apoplastic ascorbate decreased among the three types of plants, but was lowest in the sense plants. The hydrogen peroxide contents in the symplastic and apoplastic spaces were higher in sense plants, progressively lower in the wild type, followed by the antisense plants. The Arabidopsis T-DNA inserted mutant exhibited very low ascorbate oxidase activity, and its phenotype was similar to that of antisense tobacco plants. These results suggest that the suppressed expression of apoplastic AAO under salt-stress conditions leads to a relatively low level of hydrogen peroxide accumulation and a high redox state of symplastic and apoplastic ascorbate which, in turn, permits a higher seed yield.

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