Galactinol Synthase Genes Research Articles

Responses of Populus trichocarpa galactinol synthase genes to abiotic stresses

Galactinol synthase (GolS; EC 2.4.1.123) is a member of the glycosyltransferase eight family that catalyzes the first step in the biosynthesis pathway of the raffinose family of oligosaccharides (RFOs). The accumulation of RFOs in response to abiotic stress indicates a role for RFOs in stress adaptation. To obtain information on the roles of RFOs in abiotic stress adaptation in trees, we investigated the expression patterns of nine Populus trichocarpa GolS (PtrGolS) genes with special reference to stress responses. PtrGolS genes were differentially expressed in different organs, and the expressions of PtrGolS4 and PtrGolS6 were relatively high in all tested organs. The expression levels of all PtrGolS genes, except PtrGolS9, changed in response to abiotic stress in gene- and stress-type-specific manners. Moreover, short- and long-term stress treatments revealed that induction of PtrGolS by salt stress is obvious only in the early period of treatment (within 24 h), whereas water-deficit stress treatments continued to upregulate PtrGolS gene expression after two days of treatment, in addition to induction within 24 h of treatment. Consistent with these expression patterns, the galactinol content in leaves increased after four days of drought stress, but not under salt stress. Our findings suggest divergent roles for PtrGolS genes in abiotic stress responses in poplars.Electronic supplementary materialThe online version of this article (doi:10.1007/s10265-013-0597-8) contains supplementary material, which is available to authorized users.

Winter induction of the galactinol synthase gene is associated with endodormancy in chestnut trees

From the start of autumn and throughout winter dormancy, starch stored in the stems and buds of woody plants is broken down into oligosaccharides, mainly raffinose family oligosaccharides (RFOs). Given that RFO synthesis protects plants against winter cold, the enzyme galactinol synthase (catalyzing the first step in RFO synthesis) could play an important role in this process. Here, we characterized a galactinol synthase gene (CsGolS1) in chestnut and examined its expression in relation to tree dormancy. In chestnut branches, CsGolS1 mRNA levels paralleled the induction and course of winter dormancy, starting to rise at the beginning of autumn and elevated levels persisting until the onset of spring. In addition, exposure of chestnut plantlets to 4 °C induced the expression of the CsGolS1 gene in the stems. However, the permanent induction of stem CsGolS1 during winter months, rather than being a direct response to the cold, was found to be associated with the endodormancy state. This observation points to transcriptional mechanisms controlling the maintenance of endodormancy.

A cold responsive galactinol synthase gene from Medicago falcata (MfGolS1) is induced by myo‐inositol and confers multiple tolerances to abiotic stresses

Galactinol synthase (GolS, EC 2.4.1.123) catalyzes formation of galactinol and the subsequent synthesis of raffinose family oligosaccharides. The relationship of GolS to drought and salt tolerance has been well documented, however, little information is available about the role of GolS gene in cold tolerance. A coding sequence of MfGolS1 cDNA was cloned from Medicago sativa spp falcata (i.e. M. falcata), a species that exhibits greater cold tolerance than alfalfa (M. sativa). MfGolS1 transcript was not detected in untreated vegetative tissues using RNA blot hybridization; however, it was greatly induced in leaves, but not in stem and petiole, after cold treatment. Higher levels of MfGolS1 transcript were induced and maintained in M. falcata than in M. sativa during cold acclimation. Accordingly, more sugars including sucrose, galactinol, raffinose and stachyose were accumulated in M. falcata than in M. sativa. The data indicated that MfGolS1 transcript and its resultant sugar accumulation were associated with the differential cold tolerance between M. falcata and M. sativa. MfGolS1 transcript was weakly induced by dehydration and salt stresses, but not responsive to abscisic acid. MfGolS1 could be induced by myo-inositol, which is proposed to participate in cold-induced MfGolS1 expression. Overexpression of MfGolS1 in tobacco resulted in elevated tolerance to freezing and chilling in transgenic plants as a result of enhanced levels of galactinol, raffinose and stachyose. Tolerance to drought and salt stresses was also increased in the transgenic tobacco plants. It is suggested that MfGolS1 plays an important role in plant tolerance to abiotic stresses.

Cloning of galactinol synthase gene from Ammopiptanthus mongolicus and its expression in transgenic Photinia serrulata plants

A cold induced galactinol synthase gene (AmGS) and its promoter sequence were identified and cloned from the cold-tolerant tree Ammopiptanthus mongolicus by using cDNA-AFLP, RACE-PCR and TAIL-PCR strategies combined with its expression pattern analysis after cold inducing treatment. Accession number of the AmGS gene in GenBank is DQ519361. The open reading frame (ORF) region of the AmGS gene is 987 nucleotides encoding for 328 amino acid residues and a stop codon. The genomic DNA sequence of AmGS gene contains 3 exons and 2 introns. Moreover, a variety of temporal gene expression patterns of AmGS was detected, which revealed the up-regulation of AmGS gene in stresses of cold, ABA and others. Then the AmGS gene was transformed into Photinia serrulata tree by Agrobacterium-mediated transformation, and the transgenic plants exhibited higher cold-tolerance comparing with non-transformed plants.

Molecular characterization and expression of three galactinol synthase genes that confer stress tolerance in Salvia miltiorrhiza

To adapt to changes in their growing environment, plants express several stress-responsive genes. For example, the products of galactinol synthase (Gols) genes play a key role in regulating the levels of raffinose family oligosaccharides and conferring resistance to stress. We cloned and characterized three Gols genes in Salvia miltiorrhiza. Their expression followed three distinct patterns. Compared with the control, SmGols1 was up-regulated by temperature changes but was suppressed by exposure to methyl jasmonate or short-term drought. This gene had the greatest abundance of transcripts and was assigned a general function of carbon storage. SmGols2 responded to all stress and hormone treatments, and transcripts were maintained at a high level. Finally, expression of SmGols3 was weaker than the other two genes, but was increased significantly under different treatments. Over the experimental period, its expression declined to normal levels in response to all treatments except exposure to 100μM ABA, long-term drought, heat (42°C), or chilling (8°C). Based on our finding of cis-elements in the 5′ flanking regions, we concluded that these genes seem to be regulated by several HSF transcription factors. We also targeted their 90-bp conserved sequences and used them for RNA interference analysis. Some were knocked down to various extents in our transgenic lines. Fluctuations in their malondialdehyde contents under different stress treatments, as well as the rate of water loss in transformed plants, suggested that lipid peroxidation was more likely to occur in the transgenics than in the control. These results indicate that SmGols genes could have a main function in responding to cold or heat. Therefore, we believe that it is important to investigate this mechanism for tolerance in S. miltiorrhiza and to examine how expression of these SmGols and other homologs are influenced by abiotic stresses.

Identification and Functional Characterization of the GALACTINOL SYNTHASE (MoGolS1) Gene in Melissa officinalis Plants

Galactinol and rafinose accumulation in plants is associated with stressful environmental conditions such as cold, heat, or dehydration by the action of galactinols synthase (GolS) in the raffinose family of oligosaccharides biosynthetic pathway from UDP-galactose. Moreover, several reports mentioned that GolS transcription is up regulated by various environmental stresses like cold, heat, dehydration. Therefore, to determine whether MoGolS1 was induced with the abiotic stress we analyzed the expression pattern of the gene under various abiotic stresses like heat, cold, abscisic acid, sucrose and salt concentration in the lemon balm plants grown in standard MS medium. The MoGolS1 gene was 981-bp in length encoding 326 amino acids in its sequence and shared 77 and 76% sequence similarity with Arabidopsis thaliana galactinol synthase4 (AtGolS4) and AtGolS1 genes respectively. The MoGolS1 gene was strongly expressed by the abiotic stress induced by sucrose, ABA or heat shock. It was also expressed in responses to cold, Identification and Functional Characterization of the GALACTINOL SYNTHASgene induction with various stresses may be possible for itscrucial function in abiotic stress tolerance in plants, providing a good engineering target for genetic engineering.

Expression of a GALACTINOL SYNTHASE gene is positively associated with desiccation tolerance of Brassica napus seeds during development

Desiccation tolerance of seeds is positively correlated with raffinose family oligosaccharides (RFOs). However, RFOs' role in desiccation tolerance is still a matter of controversy. The aim of this work was to monitor the accumulation of RFO during acquisition of desiccation tolerance in rapeseed (Brassica napus L.). Rapeseeds become desiccation tolerant at 21-24d after flowering (DAF), and the time was coincident with an accumulation of raffinose and stachyose. A gene encoding galactinol synthase (GolS; EC2.4.1.123), involved in RFO biosynthesis, was cloned and functionally characterized. Enzymatic properties of recombinant galactinol synthase were also determined. Accumulation of BnGOLS-1 mRNA in developing rapeseeds was concomitant with dry weight deposition and the acquisition of desiccation tolerance, and was concurrent with the formation of raffinose and stachyose. The physiological implications of BnGOLS-1 expression patterns in developing seeds are discussed in light of the hypothesized role of RFOs in seed desiccation tolerance.

Jasmonate-dependent expression of a galactinol synthase gene is involved in priming of systemic fungal resistance in Arabidopsis thaliana

Previously, root colonization by the rhizobacterium, Pseudomonas chlororaphis O6, was shown to induce expression of galactinol synthase conferring systemic resistance against a fungal pathogen in cucumber leaves. Here, the Arabidopsis – Botrytis cinerea system is introduced to better understand signal transduction of galactinol and (or) raffinose family oligosaccharides (RFO) during O6-mediated induced systemic resistance (ISR). Among the 10 Arabidopsis galactinol synthase genes, only AtGolS1 was specifically induced upon infection with the fungal pathogen B. cinerea. AtGolS1 was primed by O6 colonization against the pathogen in Arabidopsis leaves. Arabidopsis T-DNA insertion mutants at the AtGolS1 gene site compromised O6-mediated ISR against the pathogen, thereby suggesting that AtGolS1 plays an important role in ISR. O6 colonization increased AtGolS1 transcription as well as ISR in several Arabidopsis signaling mutants, but not in the jar1-1 and coi1 mutant lines. Exogenous jasmonate treatment induced transcription of AtGolS1 in wild-type Col-0 plants, but salicylic acid and 1-aminocyclopropane-1-carboxylate did not. These studies on signaling mutants and target gene expression indicate that expression of AtGolS1 in response to O6 colonization is mediated through the jasmonate-dependent pathway, stimulating ISR in Arabidopsis against B. cinerea infection.

Functional Analysis of an Arabidopsis thaliana Abiotic Stress-inducible Facilitated Diffusion Transporter for Monosaccharides

Sugars play indispensable roles in biological reactions and are distributed into various tissues or organelles via transporters in plants. Under abiotic stress conditions, plants accumulate sugars as a means to increase stress tolerance. Here, we report an abiotic stress-inducible transporter for monosaccharides from Arabidopsis thaliana that is termed ESL1 (ERD six-like 1). Expression of ESL1 was induced under drought and high salinity conditions and with exogenous application of abscisic acid. Promoter analyses using beta-glucuronidase and green fluorescent protein reporters revealed that ESL1 is mainly expressed in pericycle and xylem parenchyma cells. The fluorescence of ESL1-green fluorescent protein-fused protein was detected at tonoplast in transgenic Arabidopsis plants and tobacco BY-2 cells. Furthermore, alanine-scanning mutagenesis revealed that an N-terminal LXXXLL motif in ESL1 was essential for its localization at the tonoplast. Transgenic BY-2 cells expressing mutated ESL1, which was localized at the plasma membrane, showed an uptake ability for monosaccharides. Moreover, the value of K(m) for glucose uptake activity of mutated ESL1 in the transgenic BY-2 cells was extraordinarily high, and the transport activity was independent from a proton gradient. These results indicate that ESL1 is a low affinity facilitated diffusion transporter. Finally, we detected that vacuolar invertase activity was increased under abiotic stress conditions, and the expression patterns of vacuolar invertase genes were similar to that of ESL1. Under abiotic stress conditions, ESL1 might function coordinately with the vacuolar invertase to regulate osmotic pressure by affecting the accumulation of sugar in plant cells.

Down-regulation of galactinol synthesis in oilseedBrassica napusleads to significant reduction of antinutritional oligosaccharidesThis paper is one of a selection of papers published in a Special Issue from the National Research Council of Canada – Plant Biotechnology Institute.

The utility of defatted seed meal from many crops such as canola ( Brassica napus L.) is limited by the presence of antinutritional factors, including sucrose galactosides, raffinose, and stachyose. Anaerobic breakdown of these sugars in the digestive tract of livestock is a major source of production of farm gases. In this report, the gene encoding galactinol synthase was isolated from B. napus and reintroduced into the same species in an antisense orientation to limit the production of galactinol, an important intermediate in the biosynthesis of raffinose and stachyose. This approach substantially reduced the accumulation of galactinol and stachyose in mature transgenic canola seed. Substantial changes in the mRNA levels of galactinol synthase and several sugar-related genes were also observed.

Galactinol Is a Signaling Component of the Induced Systemic Resistance Caused by Pseudomonas chlororaphis O6 Root Colonization

Root colonization by Pseudomonas chlororaphis O6 in cucumber elicited an induced systemic resistance (ISR) against Corynespora cassiicola. In order to gain insight into O6-mediated ISR, a suppressive subtractive hybridization technique was applied and resulted in the isolation of a cucumber galactinol synthase (CsGolS1) gene. The transcriptional level of CsGolS1 and the resultant galactinol content showed an increase several hours earlier under O6 treatment than in the water control plants following C. cassiicola challenge, whereas no difference was detected in the plants without a pathogen challenge. The CsGolS1-overexpressing transgenic tobacco plants demonstrated constitutive resistance against the pathogens Botrytis cinerea and Erwinia carotovora, and they also showed an increased accumulation in galactinol content. Pharmaceutical application of galactinol enhanced the resistance against pathogen infection and stimulated the accumulation of defense-related gene transcripts such as PR1a, PR1b, and NtACS1 in wild-type tobacco plants. Both the CsGolS1-overexpressing transgenic plants and the galactinol-treated wild-type tobacco plants also demonstrated an increased tolerance to drought and high salinity stresses.

The role of raffinose in the cold acclimation response of Arabidopsis thaliana

In many plants raffinose family oligosaccharides are accumulated during cold acclimation. The contribution of raffinose accumulation to freezing tolerance is not clear. Here, we investigated whether synthesis of raffinose is an essential component for acquiring frost tolerance. We created transgenic lines of Arabidopsis thaliana accessions Columbia-0 and Cape Verde Islands constitutively overexpressing a galactinol synthase (GS) gene from cucumber. GS overexpressing lines contained up to 20 times as much raffinose as the respective wild-type under non-acclimated conditions and up to 2.3 times more after 14 days of cold acclimation at 4 °C. Furthermore, we used a mutant carrying a knockout of the endogenous raffinose synthase (RS) gene. Raffinose was completely absent in this mutant. However, neither the freezing tolerance of non-acclimated leaves, nor their ability to cold acclimate were influenced in the RS mutant or in the GS overexpressing lines. We conclude that raffinose is not essential for basic freezing tolerance or for cold acclimation of A. thaliana.

Expression of the maize GALACTINOL SYNTHASE gene family: (II) Kernel abscission, environmental stress and myo-inositol influences accumulation of transcript in developing seeds and callus cells

Accumulation of transcript from three maize (Zea mays L. inbred line B73) GALACTINOL SYNTHASE (ZmGOLS) genes in abscised developing seeds and callus before and after stress was examined. Previous work showed that transcript from ZmGOLS3 predominates during seed development, whereas ZmGOLS2 transcript accumulates when triggered by certain stresses following imbibition. Abscission stimulated accumulation of ZmGOLS transcript in immature kernels. This accumulation was enhanced by increasing durations of dehydration stress. In callus tissue, neither ZmGOLS1 nor ZmGOLS3 was detected regardless of media composition or the nature or duration of the applied stress. Accumulation of ZmGOLS2 transcript in heat- or dehydration-stressed callus was induced by exogenously supplied myo-inositol in low amounts but not by sucrose. However, abundance of transcript was a poor indicator of activity of GOLS or amounts of galactinol/raffinose regardless of type of tissue, composition of media, or imposition of stress. Neither were amounts of galactinol/raffinose tightly associated with activity of GOLS nor amounts of myo-inositol, despite the ability of exogenously supplied raffinose to maintain the growth of dehydration-stressed callus. The abundance of ZmGOLS transcript did not increase in response to exogenously supplied abscisic acid.

Galactinol Synthase Gene Expression in Melon

Raffinose family oligosaccharides (RFOs) perform several physiological functions in plants. In addition to accumulating during seed formation, raffinose and stachyose are translocated in the phloem and may accumulate in response to low temperatures, drought, or salt stress. Although the synthesis of galactinol, as mediated by galactinol synthase (GAS), is the first committed step in RFO formation, its expression patterns are poorly understood in most species. We have cloned and characterized the expression of two galactinol synthase gene family members in melon (Cucumis melo L. Cantalupensis Group). Both CmGAS1 and CmGAS2 are highly expressed in mature leaves. Galactinol synthase transcription in leaves was not upregulated by either water or low temperature stresses. Transcripts of CmGAS1 were present in developing melon seeds at a time coincident with the formation of raffinose and stachyose. Based on the GAS expression and RFO accumulation patterns, we propose that RFOs in melon function in carbon translocation and seed desiccation tolerance.

Galactinol synthase (GS): increased enzyme activity and levels of mRNA due to cold and desiccation

Galactinol synthase (GS) catalyzes the first committed step in the biosynthesis of the raffinose family oligosaccharides (RFO) and could play a key regulatory role in the carbon partitioning between sucrose and RFO in the developing seed. RFO have been proposed to be osmoprotectants when plants are exposed to environmental water deficit stresses, such as cold and desiccation, and during the developmentally induced desiccation in the late maturation stage of seed development. We present the following evidences obtained from three plant species in support of these proposed roles for GS and RFO: (a) GS activity increased in kidney bean seeds upon exposure of plants to cold; (b) GS mRNA levels increased significantly upon cold exposure in the vegetative tissues of Arabidopsis thaliana, a chill resistant species, with the transcripts diminishing upon re-exposure to room temperature. A smaller increase due to cold was also observed in the siliques with the transcripts likewise disappearing upon re-exposure of the plants to room temperature; (c) we established by protein sequence comparison that a previously unidentified gene belonging to a group of ABA-independent, desiccation stress inducible genes isolated from rice with acquired chill resistance, encodes the rice homologue of the GS gene.