Plant Growth Regulation

, Volume 65, Issue 1, pp 23–34 | Cite as

Antioxidant enzymatic protection during tobacco leaf ageing is affected by cytokinin depletion

  • Zuzana Mýtinová
  • Václav Motyka
  • Daniel Haisel
  • Zuzana Lubovská
  • Alena Trávníčková
  • Petre Dobrev
  • Josef Holík
  • Naďa Wilhelmová
Original paper

Abstract

Plant ageing and senescence are associated with increased levels of reactive oxygen species. Level of cytokinins, the apparent inhibitors of plant senescence, is controlled by their irreversible degradation catalysed by cytokinin oxidase/dehydrogenase (CKX). We investigated the CKX activity, cytokinin concentration, and activities of antioxidative enzymes in tobacco (Nicotiana tabacum L. cv. Samsun NN) overexpressing the Arabidopsis gene for AtCKX2, targeted for extracellular secretion pathway. The control and AtCKX2 plants differed substantially in their phenotypes. When the lowest leaves in controls became yellow all leaves in AtCKX2 tobacco still remained green. Activities of antioxidant enzymes decreased with leaf age in both tobacco plants except for ascorbate peroxidase (APX) in the old leaves and glutathione reductase (GR) in young leaves. Enhancement of GR activity at all leaf stages, an increase of superoxide dismutase and a decline of catalase in young leaves, as well as an increase of APX in the oldest leaves were observed in AtCKX2 plant compared to control. Similar changes were detected after determination of isoenzymes on zymograms. It is evident that AtCKX2 plants had postponed onset of senescence despite the significantly lowered level of cytokinins. Enhanced antioxidant protection, especially in the oldest leaves, could subsidise this phenomenon.

Keywords

Ageing Antioxidant enzymes Cytokinins Cytokinin oxidase/dehydrogenase Tobacco 

Abbreviations

ABA

Abscisic acid

APX

Ascorbate peroxidase

AtCKX2

Transgenic tobacco overexpressing the Arabidopsis gene for CKX2

CAT

Catalase

cisZ

cis-zeatin

cisZ7G

cis-zeatin-7-glucoside

CKX

Cytokinin oxidase/dehydrogenase

DCPIP

2,6-dichlorophenolindophenol

DTT

Dithiotreitol

EDTA

Ethylendiaminetetraacetic acid

GR

Glutathione reductase

GSSG

Oxidised glutathione

iP

N6-(Δ2-isopentenyl)adenine

iP7G

N6-(Δ2-isopentenyl)adenine-7-glucoside

MTT

3(4,5-dimethyl-2-thiazolyl)-2,5diphenyl-2H-tetrazolium bromide

NBT

3,3′-(3,3′-dimethoxy-4,4′-biphenylene)bis[2-(4-nitrophenyl)-5-phenyl-2H-tetrazolium chloride]

PAR

Photosynthetically active radiation

ROS

Reactive oxygen species

SOD

Superoxide dismutase

TEMED

N,N,N′,N′-tetramethylethylenediamine

XTT

2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide, sodium salt

Z

trans-zeatin

Z7G

trans-zeatin-7-glucoside

Notes

Acknowledgments

This work was supported by the Grant Agency of the Czech Republic (522/03/0312), the Grant Agency of the Academy of Sciences of the Czech Republic (IAA600380701), the Ministry of Education, Youth and Sports CR (LC 06034) and AVOZ50380511. The authors thank Dr. Miroslav Kamínek for critical reading of manuscript and Ms. Marie Korecká and Ms. Lenka Kolčabová for excellent technical assistance.

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Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Zuzana Mýtinová
    • 2
  • Václav Motyka
    • 3
  • Daniel Haisel
    • 2
  • Zuzana Lubovská
    • 1
    • 2
  • Alena Trávníčková
    • 3
  • Petre Dobrev
    • 3
  • Josef Holík
    • 4
  • Naďa Wilhelmová
    • 2
  1. 1.Faculty of ScienceCharles UniversityPraha 2Czech Republic
  2. 2.Academy of Sciences of the Czech Republic, v.v.i.Institute of Experimental BotanyPraha 6Czech Republic
  3. 3.Academy of Sciences of the Czech RepublicInstitute of Experimental BotanyPraha 6Czech Republic
  4. 4.Academy of Sciences of the Czech RepublicInstitute of Experimental BotanyPraha 4Czech Republic

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