Expression of an antisense Datura stramonium S-adenosylmethionine decarboxylase cDNA in tobacco: Changes in enzyme activity, putrescine-spermidine ratio, rhizogenic potential, and response to methyl jasmonate

Dipartimento di Biologia e.s., University of Bologna, Via Irnerio 42, 40126 Bologna, Italy.
Journal of Plant Physiology (Impact Factor: 2.56). 06/2005; 162(5):559-71. DOI: 10.1016/j.jplph.2004.10.008
Source: PubMed


S-adenosylmethionine decarboxylase activity (SAMDC; EC leads to spermidine and spermine synthesis through specific synthases which use putrescine, spermidine and decarboxylated S-adenosylmethionine as substrates. In order to better understand the regulation of polyamine (PA), namely spermidine and spermine, biosynthesis, a SAMDC cDNA of Datura stramonium was introduced in tobacco (Nicotiana tabacum L. cv. Xanthi) in antisense orientation under the CaMV 35S promoter, by means of Agrobacterium tumefaciens and leaf disc transformation. The effect of the genetic manipulation on PA metabolism, ethylene production and plant morphology was analysed in primary transformants (R0), and in the transgenic progeny (second generation, R1) of self-fertilised primary transformants, relative to empty vector-transformed (pBin19) and wild-type (WT) controls. All were maintained in vitro by micropropagation. Primary transformants, which were confirmed by Southern and northern analyses, efficiently transcribed the antisense SAMDC gene, but SAMDC activity and PA titres did not change. By contrast, in most transgenic R1 shoots, SAMDC activity was remarkably lower than in controls, and the putrescine-to-spermidine ratio was altered, mainly due to increased putrescine, even though putrescine oxidising activity (diamine oxidase, EC did not change relative to controls. Despite the reduction in SAMDC activity, the production of ethylene, which shares with PAs the common precursor SAM, was not influenced by the foreign gene. Some plants were transferred to pots and acclimatised in a growth chamber. In these in vivo-grown second generation transgenic plants, at the vegetative stage, SAMDC activity was scarcely reduced, and PA titres did not change. Finally, the rhizogenic potential of in vitro-cultured leaf explants excised from antisense plants was significantly diminished as compared with WT ones, and the response to methyl jasmonate, a stress-mimicking compound, in terms of PA conjugation, was higher and differentially affected in transgenic leaf discs relative to WT ones. The effects of SAMDC manipulation are discussed in relation to plant generation, culture conditions and response to stress.

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Available from: Patrizia Torrigiani
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    • "The over-accumulation of conjugated PAs and induction of PA oxidation mediated by jasmonates (Jas) was also reported to occur in Hycoscyamus muticus L. root cultures, tobacco leaf disks and chickpea plants (Biondi et al., 2000, 2003; Biondi and Scaramagli , 2001; Rea et al., 2002). The effect of MeJA on conjugated PA levels is exacerbated in tobacco transgenic lines expressing a SAMDC gene in antisense, which showed low SAMDC activity and a relative accumulation of Put (Torrigiani et al., 2005). "
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    • "After evaporating the HCl and resuspending the dry residue in 4% PCA, hydrolysed samples were also dansylated . Putrescine (Put), spermidine (Spd) and spermine (Spm) were separated and identified by HPLC as described by Torrigiani et al. (2005). "

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    • "After evaporating the HCl and resuspending the dry residue in 4% PCA, hydrolysed samples were also dansylated . Putrescine (Put), spermidine (Spd) and spermine (Spm) were separated and identified by HPLC as described by Torrigiani et al. (2005). "
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