Improved Performance of Transgenic Fructan-Accumulating Tobacco under Drought Stress

Pilon-Smits, Elizabeth A. H.; Ebskamp, M.J.M.; Paul, M. J.; Jeuken, Marieke; Weisbeek, Peter; Smeekens, Sjef

doi:10.1104/pp.107.1.125

Cited by 446 publications

(209 citation statements)

References 13 publications

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“…Recently proline has been considered as an osmoprotectant, with overproduction, as in the high-P clover plants, resulting in increased tolerance to osmotic and environmental stresses (Yoshiba et al, 1997). The role of proline and other solutes such as fructans (Pilon-Smits et al, 1995) in drought resistance might not necessarily be osmotic because of their relatively small contributions to the osmolality of expressed leaf sap, unless they are largely restricted to small compartments such as the cytoplasm or organelles where their presence could become osmotically highly significant.…”

Section: mentioning

confidence: 99%

Role of proline and leaf expansion rate in the recovery of stressed white clover leaves with increased phosphorus concentration

et al. 2000

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Osmotic adjustment (OA) and increased cell-wall extensibility required for expansive leaf growth are well defined components of adaptation to water stress in dry soil, which might interact with soil phosphorus (P) concentration and defoliation frequency for intensively grazed white clover in legume-based pastures. Experiments were conducted with frequently and infrequently defoliated mini-swards of white clover growing in dry soil with low and high P concentrations. The higher yielding high-P plants were able to dry the soil to greater soil water suctions ; their leaves had lower water potential values, yet they showed fewer water stress symptoms and underwent a more complete recovery from the water stress symptoms on rewatering, than the low-P plants. High-P plants had greater OA, proline concentration and leaf expansion rate. On the other hand, low-P plants showed an increased osmotic concentration when there was no change in the total solute content per unit of leaf d. wt, indicating more loss of water from the leaf tissue. The key measures that appeared to be directly associated with plant recovery over a short period following water stress were increased proline concentration and leaf expansion rate, probably resulting from increased cell-wall extensibility rather than increased production of cells for the high-P plants.Key words : osmotic adjustment, leaf expansion rate, leaf water potential, phosphorus, white clover, proline, drought tolerance, plant recovery. The susceptibility to drought of the shallow-rooted white clover plant (Trifolium repens) is widely accepted (Hart, 1987). The decrease in osmotic potential of white clover leaves with developing water stress is reported to be insufficient to maintain pressure potential and associated metabolic activities and growth, and therefore leaves wilt and die early in the development of water stress (Turner, 1990). However, in recent studies it has been observed that an increased P supply to white clover plants growing in dry soil results in increased drought tolerance (Singh et al., 1997). Plants growing in low-P soil showed severe wilting symptoms with desiccation and drying of the leaves and stolons, and some *Author for correspondence (tel j61 8 9166 4000 ; fax j61 8 9166 4066 ; e-mail dsingh!agric.wa.gov.au). eventual mortality. These effects on low-P plants were exacerbated by frequent defoliation. However, frequently defoliated plants with adequate P supply showed minimum wilting symptoms ; they maintained higher leaf water potential (Ψ leaf ) and continued to grow in very dry soil, extracting considerably more water than the P-deficient plants. Importantly, rewatering resulted in an immediate, overnight recovery of stressed leaves for the high-P plants, but not for the low-P plants.It is not known why and how these high-P plants showed greater drought tolerance and recovery compared with low-P plants. It appears that an increased P concentration in high-P plants induced some osmotic adjustments (OA) that would improve their drought...

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Section: mentioning

confidence: 99%

Role of proline and leaf expansion rate in the recovery of stressed white clover leaves with increased phosphorus concentration

et al. 2000

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“…Genes for low-affinity bacterial and plant fructosyl transferases have been used to transform non-fructan plants such as maize, tobacco and potato (Pilon-Smits et al, 1995Caimi et al 1996 ;Vijn et al, 1997). Where full data were reported, these transgenic tissues can be shown to accumulate sucrose at concentrations which are low ( 3 mM) relative to the substrate affinity of the enzyme.…”

Section: Physiological Significancementioning

confidence: 99%

Characterization of the enzymatic polymerization of 2,6‐linked fructan by leaf extracts from timothy grass (Phleum pratense)

et al. 1999

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A fructan polymerase activity was partially purified and concentrated by sequential acid and salt precipitation from extracts of excised, illuminated leaves of timothy grass (Phleum pratense). The polymerase catalysed the de novo synthesis of oligo-and polyfructan from sucrose as sole substrate at near-physiological rates (0n5-0n9 mg g −" fresh mass h −" ; 0n9-1n5 nkat g −" ). Rates of in vitro polymerisation were high, at up to 4n1 mg cm −$ h −" (7n1 nkat cm −$ ) of total products of degree of polymerization greater than 2 (DP 2). The trisaccharides 1-kestose and 6-kestose together with oligosaccharides of up to DP c. 10 were synthesized in under 2 h at 30mC. In longer incubations, ethanol-precipitable polymers of DP l c. 10-35 (1n6-5n7 kDa) were detected by anion-exchange chromatography and pulsed amperometry. When this polymeric product was used as a primer and re-incubated with fresh enzyme and sucrose, abundant polymers of up to DP l 50 (8n1 kDa) were formed. The structure of the polymeric enzyme product was compared with native fructan from timothy leaves and with standard inulin, using glycosyl-linkage analysis followed by identification of partially methylated alditol acetate derivatives by GC-MS. The deduced structure was a linear (unbranched) 2,6-linked fructose chain terminated with glucose and fructose. The linkage structures of the native and enzyme-generated polymers were identical, increasing confidence in the physiological relevance of the activity. After ultracentrifugation of tissue homogenates at 265 000 g av , the polymerase remained in the supernatant, demonstrating no tight association with particulate components. The polymerizing reaction was dependent on enzyme concentration, requiring at least 3 g fresh mass equivalent cm −$ (c. 2n7 nkat cm −$ ) for the efficient in vitro generation of fructans of DP 3. In common with other trisaccharide-synthesizing and oligofructan-glycosylating enzymes from grasses, the polymerase reaction exhibited both a maximal velocity at pH 5n0-5n5 and a low affinity for sucrose. The polymerization reaction did not saturate fully even at 1n5 M sucrose, and the concentration causing half maximal velocity (apparent K m ) was c. 560 mM. The preparation contained substantial invertase activity (1n8 mg sucrose g −" fresh mass h −" l 1n5 nkat g −" fresh mass) with a K m for sucrose hydrolysis of 5 mM. A single peak of polymerase activity with an M r of 51 kDa was recovered from size-exclusion chromatography (SEC). Invertases of M r 51 and 110 kDa were identified in the

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“…Fructans have been implicated in drought resistance in transgenic plants (Pilon Smits et al, 1995). Further investigation of fructanaccumulating liverworts could illuminate the enzymology of fructan biosynthesis.…”

Section: mentioning

confidence: 99%

“…Transgenic tobacco plants expressing bacterial levan sucrase, which introduces the capacity for fructan synthesis, appear to perform better when subject to drought or high salinity (Pilon-Smits et al, 1995). Introduction of mannitol biosynthesis into tobacco by expressing bacterial mannitol-1-phosphate dehydrogenase also improves growth when plants are exposed to high salinity (Tarczyski, Jensen & Bohnert, 1993).…”

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confidence: 99%

Carbohydrate composition and invertase activity of the leafy liverwort Porella platyphylla

1998

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The major soluble carbohydrates in the desiccation-tolerant leafy liverwort Porella platyphylla (L.) Lindb. are sucrose and a homologous series of fructans including the trisaccharide 1-kestose. Exogenous glucose and fructose (10 mol m −$ ) did not affect the composition of the soluble carbohydrate pool. Sucrose caused an increase in the fructan pool. Sucrose also inhibited photosynthetic oxygen evolution and respiration. The fructan pool was maintained in preference to sucrose during dark starvation. Low temperature and low water potential increased the fructan pool whereas desiccation increased the proportion of high molecular weight fructan. Acid invertase activity was detected in a taxonomically diverse range of liverworts but was very low or undetectable in a range of mosses. The invertase activity from P. platyphylla was partially purified by ammonium sulphate precipitation. The reaction products of the partially purified enzyme were equimolar glucose and fructose. Kestose and higher DP fructans were not detected suggesting that, at least under the assay conditions used, the enzyme does not have sucrose : sucrose fructosyl transferase activity. The pH optimum was 4n5-5 and the K m for sucrose was 1n7 mol m −$ . Pyridoxal hydrochloride (5 mol m −$ ) caused 50 % inhibition. The coexistence of sucrose and invertase suggests that either the invertase is inactive in vivo or is in a different subcellular compartment from sucrose. The pH response shows that it would have very low activity at cytosolic pH. A large acidic vacuole was detected in P. platyphylla leaf cells by neutral-red staining in which either invertase or sucrose could be sequestered. Rehydrating desiccated P. platyphylla for 10 min resulted in a 60 % loss of extractable invertase activity. By contrast, extractable malate dehydrogenase activity increased during rehydration. Rehydrating desiccated leaves caused an increase in glucose and fructose suggesting that the sucrose pool was susceptible to invertase at this time. It is suggested that the partial inactivation of invertase during rehydration minimizes sucrose hydrolysis while membrane structure and subcellular compartmentation are re-established.

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Improved Performance of Transgenic Fructan-Accumulating Tobacco under Drought Stress

Cited by 446 publications

References 13 publications

Role of proline and leaf expansion rate in the recovery of stressed white clover leaves with increased phosphorus concentration

Role of proline and leaf expansion rate in the recovery of stressed white clover leaves with increased phosphorus concentration

Characterization of the enzymatic polymerization of 2,6‐linked fructan by leaf extracts from timothy grass (Phleum pratense)

Carbohydrate composition and invertase activity of the leafy liverwort Porella platyphylla

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