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Improved drought tolerance without undesired side effects in transgenic plants producing trehalose

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Abstract

Most organisms naturally accumulating trehalose upon stress produce the sugar in a two-step process by the action of the enzymes trehalose-6-phosphate synthase (TPS) and trehalose-6-phosphate phosphatase (TPP). Transgenic plants overexpressing TPS have shown enhanced drought tolerance in spite of minute accumulation of trehalose, amounts believed to be too small to provide a protective function. However, overproduction of TPS in plants has also been found combined with pleiotropic growth aberrations. This paper describes three successful strategies to circumvent such growth defects without loosing the improved stress tolerance. First, we introduced into tobacco a double construct carrying the genes TPS1 and TPS2 (encoding TPP) from Saccharomyces cerevisiae. Both genes are regulated by an Arabidopsis RuBisCO promoter from gene AtRbcS1A giving constitutive production of both enzymes. The second strategy involved stress-induced expression by fusing the coding region of ScTPS1 downstream of the drought-inducible Arabidopsis AtRAB18 promoter. In transgenic tobacco plants harbouring genetic constructs with either ScTPS1 alone, or with ScTPS1 and ScTPS2 combined, trehalose biosynthesis was turned on only when the plants experienced stress. The third strategy involved the use of AtRbcS1A promoter together with a transit peptide in front of the coding sequence of ScTPS1, which directed the enzyme to the chloroplasts. This paper confirms that the enhanced drought tolerance depends on unknown ameliorated water retention as the initial water status is the same in control and transgenic plants and demonstrates the influence of expression of heterologous trehalose biosynthesis genes on Arabidopsis root development.

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Abbreviations

DC:

double constitutive (in genetic constructs)

DI:

double inducible (in genetic constructs)

Glc6P:

glucose-6-phosphate

Ler:

Arabidopsis thaliana, wild-type ecotype Landsberg erecta

MCS:

multiple cloning site

ori:

origin of replication

OtsA:

Escherichia coli trehalose-6-phosphate synthase

OtsB:

E. coli trehalose-6-phosphate phosphatase

RE:

restriction enzyme

RuBisCO:

Ribulose-1,5-bisphosphate carboxylase

RWC:

relative water content

Sc:

Saccharomyces cerevisiae

SC:

single constitutive (in genetic constructs)

SCTP:

single constitutive with a transit peptide (in genetic constructs)

SI:

single inducible (in genetic constructs)

Tps1:

eukaryotic trehalose-6-phosphate synthase

Tps2:

eukaryotic trehalose-6-phosphate phosphatase

T6P:

trehalose-6-phosphate

TH:

trehalase

TP:

transit peptide

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Acknowledgements

We are grateful to Dr. John Londesborough for the clones harbouring the yeast ScTPS1 and ScTPS2 genes and for technical advice and valuable discussions. Afsaneh Ahmadzadeh for assistance with vector constructs, and Gunvor Sandman and Maj-Britt Karlsson for help with the green-house and seed collections. Gunilla Swärdh at the Uppsala Transgenic Arabidopsis Facility for Arabidopsis transformation work. Parts of this study was supported by The Swedish Technical Research Council, the Swedish Research Council for Environment, Agricultural Sciences and Spatial Planning (FORMAS).

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Karim, S., Aronsson, H., Ericson, H. et al. Improved drought tolerance without undesired side effects in transgenic plants producing trehalose. Plant Mol Biol 64, 371–386 (2007). https://doi.org/10.1007/s11103-007-9159-6

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