Plant Science 161 (2001) 179–188 Induction of shikimate dehydrogenase and peroxidase in pepper (Capsicum annuum L.) seedlings in response to copper stress and its relation to lignification Jose ´ Dı ´az *, Angeles Bernal, Federico Pomar, Fuencisla Merino Depto. de Bioloxı ´a Animal, Bioloxı ´a Vexetal e Ecoloxı ´a, Uniersidade da Corun ˜a, Campus da Zapateira s /n. E-15071. A Corun ˜a, Spain Received 5 February 2001; received in revised form 28 March 2001; accepted 28 March 2001 Abstract Pepper (Capsicum annuum L.) plants growing in a nutrient solution with excess copper, showed an increase in shikimate dehydrogenase (SKDH, EC 1.1.1.25) and peroxidase (EC 1.11.1.7) activities in the hypocotyl. In the roots, peroxidase was also induced, but SKDH activity per organ was depleted rather than enhanced. Copper stress caused stunting in the plants, reflected by a decrease in the fresh weight of all the organs. In the hypocotyl, the induction of both enzymatic activities was associated with the accumulation of soluble phenolics and lignin. The two SKDH isozymes present in the control hypocotyls (SKDH-3 and SKDH-4) increased in a similar proportion after copper stress. In the case of peroxidases, two new isozymes (PRX-A2 and PRX-A4) were detected in copper-stressed hypocotyls, and the other two isoperoxidases, PRX-B and PRX-A3, were enhanced c. 10 and three times, respectively, with respect to the control. The application of the chelator EDTA was able to counteract all the stress effects of the metal cited above. The role of these enzymes in phenolic metabolism under heavy metal stress is discussed. © 2001 Elsevier Science Ireland Ltd. All rights reserved. Keywords: Capsicum annuum; Copper stress; Lignin; Peroxidase; Phenolics; Shikimate dehydrogenase www.elsevier.com/locate/plantsci 1. Introduction Heavy metals are present in trace concentrations in different compartments of the terrestrial ecosys- tems, and some of them are essential micronutri- ents for plants and other organisms [1]. However, sometimes, the presence of phytotoxic amounts of metals occurs in soils, usually produced by agricul- tural and industrial activities [2]. Plants may take up an excess of such heavy metals, which eventu- ally affect different physiological processes. Stunted growth, necrosis, leaf epinasty, chlorosis and red – brownish discoloration are characteristic symptoms visible to the naked eye in plants suffer- ing from severe metal toxicity [3], but are the consequence of the alteration of many processes at the cellular level. Copper is one of the metals that plants need as an essential nutrient, but it can turn into a prob- lem if it accumulates excessively in soils. Usual causes of high levels of copper in the soil are mining activities or the prolonged application of copper-based fungicides, such as the Bordeaux mixture, in old orchards and vineyards [2,4]. Cop- per stress interferes with plant physiology in sev- eral ways, including the inhibition of processes such as photosynthesis, respiration or nitrogen fixation [4,5]. Other effects of copper at the cellu- lar level are the alteration of membrane integrity, the formation of active oxygen species and the Abbreiations: DTT, dithiothreitol; IAA, indol acetic acid; IEF, isoelectric focusing; 2-ME, 2-mercaptoethanol; MTT, 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; PAGE, poly- acrylamide gel electrophoresis; PAL, phenylalanine ammonia lyase; PMS, phenazine methosulfate; PRX, peroxidase; PVPP, polyvinyl- polypyrrolidone; SKDH, shikimate dehydrogenase. * Corresponding author. Tel.: +34-981-167000; fax: +34-981- 167065. E-mail address: josefv@udc.es (J. Dı ´az). 0168-9452/01/$ - see front matter © 2001 Elsevier Science Ireland Ltd. All rights reserved. PII:S0168-9452(01)00410-1