Multifunctional fructans and raffinose family oligosaccharides

Galactinol synthase (GolS) is a key enzyme in the synthesis of raffinose family oligosaccharides that function as osmoprotectants in plant cells. In leaves of Arabidopsis (Arabidopsis thaliana) plants overexpressing heat shock transcription factor A2 (HsfA2), the transcription of GolS1, -2, and -4 and raffinose synthase 2 (RS2) was highly induced; thus, levels of galactinol and raffinose increased compared with those in wild-type plants under control growth conditions. In leaves of the wild-type plants, treatment with 50 mum methylviologen (MV) increased the transcript levels of not only HsfA2, but also GolS1, -2, -3, -4, and -8 and RS2, -4, -5, and -6, the total activities of GolS isoenzymes, and the levels of galactinol and raffinose. GolS1- or GolS2-overexpressing Arabidopsis plants (Ox-GolS1-11, Ox-GolS2-8, and Ox-GolS2-29) had increased levels of galactinol and raffinose in the leaves compared with wild-type plants under control growth conditions. High intracellular levels of galactinol and raffinose in the transgenic plants were correlated with increased tolerance to MV treatment and salinity or chilling stress. Galactinol and raffinose effectively protected salicylate from attack by hydroxyl radicals in vitro. These findings suggest the possibility that galactinol and raffinose scavenge hydroxyl radicals as a novel function to protect plant cells from oxidative damage caused by MV treatment, salinity, or chilling.

Sucrose cleavage is vital to multicellular plants, not only for the allocation of crucial carbon resources but also for the initiation of hexose-based sugar signals in importing structures. Only the invertase and reversible sucrose synthase reactions catalyze known paths of sucrose breakdown in vivo. The regulation of these reactions and its consequences has therefore become a central issue in plant carbon metabolism. Primary mechanisms for this regulation involve the capacity of invertases to alter sugar signals by producing glucose rather than UDPglucose, and thus also two-fold more hexoses than are produced by sucrose synthase. In addition, vacuolar sites of cleavage by invertases could allow temporal control via compartmentalization. In addition, members of the gene families encoding either invertases or sucrose synthases respond at transcriptional and posttranscriptional levels to diverse environmental signals, including endogenous changes that reflect their own action (e.g. hexoses and hexose-responsive hormone systems such as abscisic acid [ABA] signaling). At the enzyme level, sucrose synthases can be regulated by rapid changes in sub-cellular localization, phosphorylation, and carefully modulated protein turnover. In addition to transcriptional control, invertase action can also be regulated at the enzyme level by highly localized inhibitor proteins and by a system that has the potential to initiate and terminate invertase activity in vacuoles. The extent, path, and site of sucrose metabolism are thus highly responsive to both internal and external environmental signals and can, in turn, dramatically alter development and stress acclimation.

Raffinose family oligosaccharides (RFO) accumulating during seed development are thought to play a role in the desiccation tolerance of seeds. However, the functions of RFO in desiccation tolerance have not been elucidated. Here we examine the functions of RFO in Arabidopsis thaliana plants under drought- and cold-stress conditions, based on the analyses of function and expression of genes involved in RFO biosynthesis. Sugar analysis showed that drought-, high salinity- and cold-treated Arabidopsis plants accumulate a large amount of raffinose and galactinol, but not stachyose. Raffinose and galactinol were not detected in unstressed plants. This suggests that raffinose and galactinol are involved in tolerance to drought, high salinity and cold stresses. Galactinol synthase (GolS) catalyses the first step in the biosynthesis of RFO from UDP-galactose. We identified three stress-responsive GolS genes (AtGolS1, 2 and 3) among seven Arabidopsis GolS genes. AtGolS1 and 2 were induced by drought and high-salinity stresses, but not by cold stress. By contrast, AtGolS3 was induced by cold stress but not by drought or salt stress. All the GST fusion proteins of GST-AtGolS1, 2 and 3 expressed in Escherichia coli had galactinol synthase activities. Overexpression of AtGolS2 in transgenic Arabidopsis caused an increase in endogenous galactinol and raffinose, and showed reduced transpiration from leaves to improve drought tolerance. These results show that stress-inducible galactinol synthase plays a key role in the accumulation of galactinol and raffinose under abiotic stress conditions, and that galactinol and raffinose may function as osmoprotectants in drought-stress tolerance of plants.