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BIOCHEMICAL PROCESSES AND MACROMOLECULAR STRUCTURES

Ectopic Expression of Maize Polyamine Oxidase and Pea Copper Amine Oxidase in the Cell Wall of Tobacco Plants¹

Department of Biology, Università degli Studi Roma Tre, Rome 00146, Italy (G.R., V.G., P.F., R.F., R.A., P.T.); Department of Biology and Plant Pathology, University of Bari, Bari 70126, Italy (M.C.d.P., L.D.G.); Ente per le Nuove tecnologie, l'Energia e l'Ambiente, Casaccia 00060, Italy (R.T.); Biology Department, University of Bologna, Bologna 40126, Italy (S.B.); and Interdepartmental Centre for Biomedical Researches, University Campus Bio-Medico, Rome 00155, Italy (L.D.G.)

To test the feasibility of altering polyamine levels by influencing their catabolic pathway, we obtained transgenic tobacco (Nicotiana tabacum) plants constitutively expressing either maize (Zea mays) polyamine oxidase (MPAO) or pea (Pisum sativum) copper amine oxidase (PCuAO), two extracellular and H₂O₂-producing enzymes. Despite the high expression levels of the transgenes in the extracellular space, the amount of free polyamines in the homozygous transgenic plants was similar to that in the wild-type ones, suggesting either a tight regulation of polyamine levels or a different compartmentalization of the two recombinant proteins and the bulk amount of endogenous polyamines. Furthermore, no change in lignification levels and plant morphology was observed in the transgenic plants compared to untransformed plants, while a small but significant change in reactive oxygen species-scavenging capacity was verified. Both the MPAO and the PCuAO tobacco transgenic plants produced high amounts of H₂O₂ only in the presence of exogenously added enzyme substrates. These observations provided evidence for the limiting amount of freely available polyamines in the extracellular space in tobacco plants under physiological conditions, which was further confirmed for untransformed maize and pea plants. The amount of H₂O₂ produced by exogenously added polyamines in cell suspensions from the MPAO transgenic plants was sufficient to induce programmed cell death, which was sensitive to catalase treatment and required gene expression and caspase-like activity. The MPAO and PCuAO transgenic plants represent excellent tools to study polyamine secretion and conjugation in the extracellular space, as well as to determine when and how polyamine catabolism actually intervenes both in cell wall development and in response to stress.

² Present address: Institute of Crystallography, Consiglio Nazionale delle Ricerche, Monterotondo, Rome 00016, Italy.

³ Present address: Ente Tabacchi Italiano, Naples, Italy.

⁴ Present address: Department of Agrobiology and Agrochemistry, University of Tuscia, Viterbo, Italy.

Article, publication date, and citation information can be found at www.plantphysiol.org/cgi/doi/10.1104/pp.103.036764.

^* Corresponding author; e-mail tavlador{at}uniroma3.it; fax 39–0655176321.

Received December 4, 2003; returned for revision January 20, 2004; accepted January 22, 2004.

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