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Enhancement of stress tolerance in transgenic tobacco plants constitutively expressing AtIpk2beta, an inositol polyphosphate 6-/3-kinase from Arabidopsis thaliana.

Yang L, Tang R, Zhu J, Liu H, Mueller-Roeber B, Xia H, Zhang H - Plant Mol. Biol. (2007)

Bottom Line: Here we expressed Arabidopsis inositol polyphosphate 6-/3-kinase (AtIpk2beta) in two heterologous systems, i.e. the yeast Saccharomyces cerevisiae and in tobacco (Nicotiana tabacum), and tested the effect on abiotic stress tolerance.Transgenic tobacco plants constitutively expressing AtIpk2beta under the control of the Cauliflower Mosaic Virus 35S promoter were generated and found to exhibit improved tolerance to diverse abiotic stresses when compared to wild type plants.Expression patterns of various stress responsive genes were enhanced, and the activities of anti-oxidative enzymes were elevated in transgenic plants, suggesting a possible involvement of AtIpk2beta in plant stress responses.

View Article: PubMed Central - PubMed

Affiliation: National Key Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 300 Fenglin Road, Shanghai 200032, China.

ABSTRACT
Inositol phosphates (IPs) and their turnover products have been implicated to play important roles in stress signaling in eukaryotic cells. In higher plants genes encoding inositol polyphosphate kinases have been identified previously, but their physiological functions have not been fully resolved. Here we expressed Arabidopsis inositol polyphosphate 6-/3-kinase (AtIpk2beta) in two heterologous systems, i.e. the yeast Saccharomyces cerevisiae and in tobacco (Nicotiana tabacum), and tested the effect on abiotic stress tolerance. Expression of AtIpk2beta rescued the salt-, osmotic- and temperature-sensitive growth defects of a yeast mutant strain (arg82Delta) that lacks inositol polyphosphate multikinase activity encoded by the ARG82/IPK2 gene. Transgenic tobacco plants constitutively expressing AtIpk2beta under the control of the Cauliflower Mosaic Virus 35S promoter were generated and found to exhibit improved tolerance to diverse abiotic stresses when compared to wild type plants. Expression patterns of various stress responsive genes were enhanced, and the activities of anti-oxidative enzymes were elevated in transgenic plants, suggesting a possible involvement of AtIpk2beta in plant stress responses.

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Germination rate and early seedling development of transgenic plants. (a–c) Percentage of germinating seeds of wild type (WT) and transgenic T2 plants (S13, S15) grown on MS medium supplemented with 0-, 100- or 200-mM NaCl. Results are presented as means and standard errors from three independent experiments (≥100 seeds of each line were sown for each experiment). (d) Wild type (WT) and transgenic seeds (S13, S15) geminated on MS medium supplemented with 50 or 150-mM NaCl. Photo was taken three weeks after seeds were sown. (e) Wild type (WT) and transgenic lines (S13, S14, S15) grown on MS medium supplemented with different concentrations of NaCl for 2 months. (f) Root bending assay of wild type (WT) and transgenic plants (S13, S15) on MS medium supplemented with 0 or 150-mM NaCl. Representative pictures are shown. (g) Primary root lengths of wild type (WT) and transgenic plants (S13, S15) in the presence of different salt concentrations. Root growth under control condition (0-mM NaCl) was set to 100%. The results are shown as means and standard errors from three independent experiments (≥15 seeds of each line were sown for each experiment). *** indicates significant differences in comparison to the wild type at P < 0.001 (Student’s t-test)
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Fig3: Germination rate and early seedling development of transgenic plants. (a–c) Percentage of germinating seeds of wild type (WT) and transgenic T2 plants (S13, S15) grown on MS medium supplemented with 0-, 100- or 200-mM NaCl. Results are presented as means and standard errors from three independent experiments (≥100 seeds of each line were sown for each experiment). (d) Wild type (WT) and transgenic seeds (S13, S15) geminated on MS medium supplemented with 50 or 150-mM NaCl. Photo was taken three weeks after seeds were sown. (e) Wild type (WT) and transgenic lines (S13, S14, S15) grown on MS medium supplemented with different concentrations of NaCl for 2 months. (f) Root bending assay of wild type (WT) and transgenic plants (S13, S15) on MS medium supplemented with 0 or 150-mM NaCl. Representative pictures are shown. (g) Primary root lengths of wild type (WT) and transgenic plants (S13, S15) in the presence of different salt concentrations. Root growth under control condition (0-mM NaCl) was set to 100%. The results are shown as means and standard errors from three independent experiments (≥15 seeds of each line were sown for each experiment). *** indicates significant differences in comparison to the wild type at P < 0.001 (Student’s t-test)

Mentions: We first studied the salt tolerance of AtIpk2β expressing plants during germination and early stages of seedling development. Seeds of wild type and the three homozygous transgenic lines S13, S14 and S15 were germinated on MS medium containing different concentrations of NaCl. Germination rates of wild type and transgenic seeds were similar when cultured on normal MS medium (Fig. 3a). However, germination of wild type seeds was significantly impaired on MS medium supplemented with 100- or 200-mM NaCl, whereas germination of transgenic seeds was less strongly affected. After eight days on high salt MS medium (100-mM NaCl), only ∼28% of the wild type seeds, but more than 70% of the seeds obtained from AtIpk2β expressing plants germinated (Fig. 3b). When sown on MS medium supplemented with 200-mM NaCl, only ∼7% of the wild type seeds, but ∼30% of the transgenic seeds germinated after 18 days (Fig. 3c). At low salt concentration (50-mM NaCl), wild type and transgenic seeds displayed similar germination rates (data now shown). At 150-mM NaCl, both wild type and transgenic seeds germinated. However, development of wild type seedlings was impaired; plants produced smaller leaves and less root biomass than did transgenic seedlings (Fig. 3d). To further test the effect of salt on growth, 15-day-old wild type and transgenic seedlings were transferred to MS medium supplemented with NaCl at different concentrations. After two additional months, strong growth retardation was observed in wild type seedlings. Inhibition of growth was less evident in transgenic lines (Fig. 3e). A root-bending assay has previously been used to test the effect of elevated salt concentration on root growth (An et al. 2007; Zhu et al. 1998). At 50-mM NaCl on MS medium, root growth of both wild type and transgenic lines was accelerated, but it was inhibited at increasing NaCl concentrations. At 150-mM NaCl, inhibition of root growth was significantly more pronounced in wild type than in transgenic plants (Fig. 3f, g).Fig. 3


Enhancement of stress tolerance in transgenic tobacco plants constitutively expressing AtIpk2beta, an inositol polyphosphate 6-/3-kinase from Arabidopsis thaliana.

Yang L, Tang R, Zhu J, Liu H, Mueller-Roeber B, Xia H, Zhang H - Plant Mol. Biol. (2007)

Germination rate and early seedling development of transgenic plants. (a–c) Percentage of germinating seeds of wild type (WT) and transgenic T2 plants (S13, S15) grown on MS medium supplemented with 0-, 100- or 200-mM NaCl. Results are presented as means and standard errors from three independent experiments (≥100 seeds of each line were sown for each experiment). (d) Wild type (WT) and transgenic seeds (S13, S15) geminated on MS medium supplemented with 50 or 150-mM NaCl. Photo was taken three weeks after seeds were sown. (e) Wild type (WT) and transgenic lines (S13, S14, S15) grown on MS medium supplemented with different concentrations of NaCl for 2 months. (f) Root bending assay of wild type (WT) and transgenic plants (S13, S15) on MS medium supplemented with 0 or 150-mM NaCl. Representative pictures are shown. (g) Primary root lengths of wild type (WT) and transgenic plants (S13, S15) in the presence of different salt concentrations. Root growth under control condition (0-mM NaCl) was set to 100%. The results are shown as means and standard errors from three independent experiments (≥15 seeds of each line were sown for each experiment). *** indicates significant differences in comparison to the wild type at P < 0.001 (Student’s t-test)
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Fig3: Germination rate and early seedling development of transgenic plants. (a–c) Percentage of germinating seeds of wild type (WT) and transgenic T2 plants (S13, S15) grown on MS medium supplemented with 0-, 100- or 200-mM NaCl. Results are presented as means and standard errors from three independent experiments (≥100 seeds of each line were sown for each experiment). (d) Wild type (WT) and transgenic seeds (S13, S15) geminated on MS medium supplemented with 50 or 150-mM NaCl. Photo was taken three weeks after seeds were sown. (e) Wild type (WT) and transgenic lines (S13, S14, S15) grown on MS medium supplemented with different concentrations of NaCl for 2 months. (f) Root bending assay of wild type (WT) and transgenic plants (S13, S15) on MS medium supplemented with 0 or 150-mM NaCl. Representative pictures are shown. (g) Primary root lengths of wild type (WT) and transgenic plants (S13, S15) in the presence of different salt concentrations. Root growth under control condition (0-mM NaCl) was set to 100%. The results are shown as means and standard errors from three independent experiments (≥15 seeds of each line were sown for each experiment). *** indicates significant differences in comparison to the wild type at P < 0.001 (Student’s t-test)
Mentions: We first studied the salt tolerance of AtIpk2β expressing plants during germination and early stages of seedling development. Seeds of wild type and the three homozygous transgenic lines S13, S14 and S15 were germinated on MS medium containing different concentrations of NaCl. Germination rates of wild type and transgenic seeds were similar when cultured on normal MS medium (Fig. 3a). However, germination of wild type seeds was significantly impaired on MS medium supplemented with 100- or 200-mM NaCl, whereas germination of transgenic seeds was less strongly affected. After eight days on high salt MS medium (100-mM NaCl), only ∼28% of the wild type seeds, but more than 70% of the seeds obtained from AtIpk2β expressing plants germinated (Fig. 3b). When sown on MS medium supplemented with 200-mM NaCl, only ∼7% of the wild type seeds, but ∼30% of the transgenic seeds germinated after 18 days (Fig. 3c). At low salt concentration (50-mM NaCl), wild type and transgenic seeds displayed similar germination rates (data now shown). At 150-mM NaCl, both wild type and transgenic seeds germinated. However, development of wild type seedlings was impaired; plants produced smaller leaves and less root biomass than did transgenic seedlings (Fig. 3d). To further test the effect of salt on growth, 15-day-old wild type and transgenic seedlings were transferred to MS medium supplemented with NaCl at different concentrations. After two additional months, strong growth retardation was observed in wild type seedlings. Inhibition of growth was less evident in transgenic lines (Fig. 3e). A root-bending assay has previously been used to test the effect of elevated salt concentration on root growth (An et al. 2007; Zhu et al. 1998). At 50-mM NaCl on MS medium, root growth of both wild type and transgenic lines was accelerated, but it was inhibited at increasing NaCl concentrations. At 150-mM NaCl, inhibition of root growth was significantly more pronounced in wild type than in transgenic plants (Fig. 3f, g).Fig. 3

Bottom Line: Here we expressed Arabidopsis inositol polyphosphate 6-/3-kinase (AtIpk2beta) in two heterologous systems, i.e. the yeast Saccharomyces cerevisiae and in tobacco (Nicotiana tabacum), and tested the effect on abiotic stress tolerance.Transgenic tobacco plants constitutively expressing AtIpk2beta under the control of the Cauliflower Mosaic Virus 35S promoter were generated and found to exhibit improved tolerance to diverse abiotic stresses when compared to wild type plants.Expression patterns of various stress responsive genes were enhanced, and the activities of anti-oxidative enzymes were elevated in transgenic plants, suggesting a possible involvement of AtIpk2beta in plant stress responses.

View Article: PubMed Central - PubMed

Affiliation: National Key Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 300 Fenglin Road, Shanghai 200032, China.

ABSTRACT
Inositol phosphates (IPs) and their turnover products have been implicated to play important roles in stress signaling in eukaryotic cells. In higher plants genes encoding inositol polyphosphate kinases have been identified previously, but their physiological functions have not been fully resolved. Here we expressed Arabidopsis inositol polyphosphate 6-/3-kinase (AtIpk2beta) in two heterologous systems, i.e. the yeast Saccharomyces cerevisiae and in tobacco (Nicotiana tabacum), and tested the effect on abiotic stress tolerance. Expression of AtIpk2beta rescued the salt-, osmotic- and temperature-sensitive growth defects of a yeast mutant strain (arg82Delta) that lacks inositol polyphosphate multikinase activity encoded by the ARG82/IPK2 gene. Transgenic tobacco plants constitutively expressing AtIpk2beta under the control of the Cauliflower Mosaic Virus 35S promoter were generated and found to exhibit improved tolerance to diverse abiotic stresses when compared to wild type plants. Expression patterns of various stress responsive genes were enhanced, and the activities of anti-oxidative enzymes were elevated in transgenic plants, suggesting a possible involvement of AtIpk2beta in plant stress responses.

Show MeSH
Related in: MedlinePlus