Molecular cloning, characteristics and low temperature response of raffinose synthase gene in Cucumis sativus L.

Sucrose synthase (SUS; EC 2.4.1.13) plays important roles in sugar metabolism and abiotic stress response. But the genes encoding SUS in cucumber (Cucumis sativus L.) have not been well studied. Here, we isolated four cucumber sucrose synthase genes (CsSUS). Among them, CsSUS3, which highly expressed in the roots, was chosen for further study. Immunolocalization and subcellular localization analysis indicated that CsSUS3 localized in the cytosol and the plasma membrane, and mainly existed in the companion cells of phloem in the roots. When suffering hypoxia stress from flooding, CsSUS3 expression and SUS activity in roots increased, especially in the lateral roots; moreover, the soluble SUS activity increased clearly, but the membrane fraction hardly changed. Compared with the wild-type cucumbers, the transgenic lines with antisense expression of CsSUS3 were more sensitive to flooding. After 6 d of flooding, the SUS activity, soluble sugar and uridine 5′-diphosphate glucose (UDPG) content and the ratio of ATP/ADP in the roots of transgenic plants were significantly lower than that in wildtype plants. Moreover, the transgenic lines grew more slowly with more yellow necrosis in the leaves. These findings suggested CsSUS3 participated in resisting hypoxic stress. Furthermore, the mechanism of CsSUS3 in resisting hypoxic stress was also discussed.

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“…25 using the cotyledon transformation method (Sui et al . ). Briefly, 2‐day‐old cotyledons were cut into two parts.…”

Section: Methodsmentioning

confidence: 99%

“…The antisense vector was transformed into cucumber cultivar Guonong No. 25 using the cotyledon transformation method (Sui et al 2012). Briefly, 2-day-old cotyledons were cut into two parts.…”

Section: Agrobacterium-mediated Cucumber Transformationmentioning

confidence: 99%

Antisense suppression of cucumber (Cucumis sativus L.) sucrose synthase 3 (CsSUS3) reduces hypoxic stress tolerance

Wang

Sui

Guo

et al. 2013

Plant Cell & Environment

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“…Both sense and antisense vectors were transformed into Agrobacterium tumefaciens strain LBA4404. The sense and antisense vectors were transformed into the cucumber pure line 'Xintaimici' using the fresh expanding cotyledon disk transformation method (Liu et al, 2010;Sui et al, 2012). Briefly, 1 d after the seeds germinated, the growing points and the upper halves of cotyledons were removed, and the other halves of cotyledons were dipped in the A. tumefaciens-diluted suspension containing one-half-strength Murashige and Skoog liquid medium (optical density at 600 nm = 0.3-0.5) for 15 min.…”

Section: Generation Of Transgenic Cucumber Plantsmentioning

confidence: 99%

Down-Regulating CsHT1, a Cucumber Pollen-Specific Hexose Transporter, Inhibits Pollen Germination, Tube Growth, and Seed Development

et al. 2015

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“…To date, the only extensive biochemical characterisation of a RS has been reported from pea seeds [7], cucumber [8] and to a lesser degree from rice [9]. Arabidopsis is reported to contain six putative abiotic stress-inducible RS genes [ RS1-6 , 10].…”

Section: Introductionmentioning

confidence: 99%

Abiotic stress-induced accumulation of raffinose in Arabidopsis leaves is mediated by a single raffinose synthase (RS5, At5g40390)

2013

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BackgroundThe sucrosylgalactoside oligosaccharide raffinose (Raf, Suc-Gal1) accumulates in Arabidopsis leaves in response to a myriad of abiotic stresses. Whilst galactinol synthases (GolS), the first committed enzyme in Raf biosynthesis are well characterised in Arabidopsis, little is known of the second biosynthetic gene/enzyme raffinose synthase (RS). Conflicting reports suggest the existence of either one or six abiotic stress-inducible RSs (RS-1 to -6) occurring in Arabidopsis. Indirect evidence points to At5g40390 being responsible for low temperature-induced Raf accumulation in Arabidopsis leaves.ResultsBy heterologously expressing At5g40390 in E.coli, we demonstrate that crude extracts synthesise Raf in vitro, contrary to empty vector controls. Using two independent loss-of-function mutants for At5g40390 (rs 5–1 and 5–2), we confirm that this RS is indeed responsible for Raf accumulation during low temperature-acclimation (4°C), as previously reported. Surprisingly, leaves of mutant plants also fail to accumulate any Raf under diverse abiotic stresses including water-deficit, high salinity, heat shock, and methyl viologen-induced oxidative stress. Correlated to the lack of Raf under these abiotic stress conditions, both mutant plants lack the typical stress-induced RafS activity increase observed in the leaves of wild-type plants.ConclusionsCollectively our findings point to a single abiotic stress-induced RS isoform (RS5, At5g40390) being responsible for Raf biosynthesis in Arabidopsis leaves. However, they do not support a single RS hypothesis since the seeds of both mutant plants still contained Raf, albeit at 0.5-fold lower concentration than seeds from wild-type plants, suggesting the existence of at least one other seed-specific RS. These results also unambiguously discount the existence of six stress-inducible RS isoforms suggested by recent reports.

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Molecular cloning, characteristics and low temperature response of raffinose synthase gene in Cucumis sativus L.

Cited by 51 publications

References 52 publications

Antisense suppression of cucumber (Cucumis sativus L.) sucrose synthase 3 (CsSUS3) reduces hypoxic stress tolerance

Antisense suppression of cucumber (Cucumis sativus L.) sucrose synthase 3 (CsSUS3) reduces hypoxic stress tolerance

Down-Regulating CsHT1, a Cucumber Pollen-Specific Hexose Transporter, Inhibits Pollen Germination, Tube Growth, and Seed Development

Abiotic stress-induced accumulation of raffinose in Arabidopsis leaves is mediated by a single raffinose synthase (RS5, At5g40390)

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