Background: Sm-like proteins are highly conserved proteins that form the core of the U6 ribonucleoprotein and function in several mRNA metabolism processes, including pre-mRNA splicing. Despite their wide occurrence in all eukaryotes, little is known about the roles of Sm-like proteins in the regulation of splicing. Results: Here, through comprehensive transcriptome analyses, we demonstrate that depletion of the Arabidopsis supersensitive to abscisic acid and drought 1 gene (SAD1), which encodes Sm-like protein 5 (LSm5), promotes an inaccurate selection of splice sites that leads to a genome-wide increase in alternative splicing. In contrast, overexpression of SAD1 strengthens the precision of splice-site recognition and globally inhibits alternative splicing. Further, SAD1 modulates the splicing of stress-responsive genes, particularly under salt-stress conditions. Finally, we find that overexpression of SAD1 in Arabidopsis improves salt tolerance in transgenic plants, which correlates with an increase in splicing accuracy and efficiency for stress-responsive genes. Conclusions: We conclude that SAD1 dynamically controls splicing efficiency and splice-site recognition in Arabidopsis, and propose that this may contribute to SAD1-mediated stress tolerance through the metabolism of transcripts expressed from stress-responsive genes. Our study not only provides novel insights into the function of Sm-like proteins in splicing, but also uncovers new means to improve splicing efficiency and to enhance stress tolerance in a higher eukaryote.

Low temperature, drought, and high salinity are common stress conditions that adversely affect plant growth and crop production. The cellular and molecular responses of plants to environmental stress have been studied intensively ([Thomashow, 1999][1]; [Hasegawa et al., 2000][2]). Understanding the

Drought, high salinity and freezing impose osmotic stress on plants. Plants respond to the stress in part by modulating gene expression, which eventually leads to the restoration of cellular homeostasis, detoxification of toxins and recovery of growth. The signal transduction pathways mediating these adaptations can be dissected by combining forward and reverse genetic approaches with molecular, biochemical and physiological studies. Arabidopsis is a useful genetic model system for this purpose and its relatives including the halophyte Thellungiella halophila, can serve as valuable complementary genetic model systems.

A large genetic screen for sos (for salt overly sensitive) mutants was performed in an attempt to isolate mutations in any gene with an sos phenotype. Our search yielded 28 new alleles of sos1, nine mutant alleles of a newly identified locus, SOS2, and one allele of a third salt tolerance locus, SOS3. The sos2 mutations, which are recessive, were mapped to the lower arm of chromosome V, ~2.3 centimorgans away from the marker PHYC. Growth measurements demonstrated that sos2 mutants are specifically hypersensitive to inhibition by Na⁺ or Li⁺ and not hypersensitive to general osmotic stresses. Interestingly, the SOS2 locus is also necessary for K⁺ nutrition because sos2 mutants were unable to grow on a culture medium with a low level of K⁺. The expression of several salt-inducible genes was superinduced in sos2 plants. The salt tolerance of sos1, sos2, and sos3 mutants correlated with their K⁺ tissue content but not their Na⁺ tissue content. Double mutant analysis indicated that the SOS genes function in the same pathway. Based on these results, a genetic model for salt tolerance mechanisms in Arabidopsis is presented in which SOS1, SOS2, and SOS3 are postulated to encode regulatory components controlling plant K⁺ nutrition that in turn is essential for salt tolerance.

Drought and high salinity induce the expression of many plant genes. To understand the signal transduction mechanisms underlying the activation of these genes, we carried out a genetic screen to isolate Arabidopsis mutants defective in osmotic stress-regulated gene induction. Here we report the isolation, characterization, and cloning of a mutation, los6, which diminished osmotic stress activation of a reporter gene. RNA blot analysis indicates that under osmotic stress the transcript levels for stress-responsive genes such as RD29A, COR15A,KIN1, COR47, RD19, andADH are lower in los6 plants than in wild type plants. los6 plants were found to have reduced phytohormone abscisic acid (ABA) accumulation and to be allelic to the ABA-deficient mutant, aba1. LOS6/ABA1 encodes a zeaxanthin epoxidase that functions in ABA biosynthesis. Its expression is enhanced by osmotic stress. Furthermore, we found that there exists a positive feedback regulation by ABA on the expression of LOS6/ABA1, which may underscore a quick adaptation strategy for plants under osmotic stress. Similar positive regulation by ABA also exists for other ABA biosynthesis genes AAO3 and LOS5/ABA3 and in certain genetic backgrounds, NCED3. This feedback regulation by ABA is impaired in the ABA-insensitive mutantabi1 but not in abi2. Moreover, the up-regulation of LOS6/ABA1, LOS5/ABA3,AAO3, and NCED3 by osmotic stress is reduced substantially in ABA-deficient mutants. Transgenic plants overexpressing LOS6/ABA1 showed an increasedRD29A-LUC expression under osmotic stress. These results suggest that the level of gene induction by osmotic stress is dependent on the dosage of the zeaxanthin epoxidase enzyme.