title: High cytokinin levels induce a hypersensitive-like response in tobacco.
abstract: OBJECTIVE
  Cytokinins are positive regulators of shoot development. However, it has previously been demonstrated that efficient activation of the cytokinin biosynthesis gene ipt can cause necrotic lesions and wilting in tobacco leaves. Some plant pathogens reportedly use their ability to produce cytokinins in disease development. In response to pathogen attacks, plants can trigger a hypersensitive response that rapidly kills cells near the infection site, depriving the pathogen of nutrients and preventing its spread. In this study, a diverse set of processes that link ipt activation to necrotic lesion formation were investigated in order to evaluate the potential of cytokinins as signals and/or mediators in plant defence against pathogens.
  
  
  METHODS
  The binary pOp-ipt/LhGR system for dexamethasone-inducible ipt expression was used to increase endogenous cytokinin levels in transgenic tobacco. Changes in the levels of cytokinins and the stress hormones salicylic, jasmonic and abscisic acid following ipt activation were determined by ultra-performance liquid chromatography-electrospray tandem mass spectrometry (UPLC-MS/MS). Trends in hydrogen peroxide content and lipid peroxidation were monitored using the potassium iodide and malondialdehyde assays. The subcellular distribution of hydrogen peroxide was investigated using 3,3'-diaminobenzidine staining. The dynamics of transcripts related to photosynthesis and pathogen response were analysed by reverse transcription followed by quantitative PCR. The effects of cytokinins on photosynthesis were deciphered by analysing changes in chlorophyll fluorescence and leaf gas exchange.
  
  
  RESULTS
  Plants can produce sufficiently high levels of cytokinins to trigger fast cell death without any intervening chlorosis - a hallmark of the hypersensitive response. The results suggest that chloroplastic hydrogen peroxide orchestrates the molecular responses underpinning the hypersensitive-like response, including the inhibition of photosynthesis, elevated levels of stress hormones, oxidative membrane damage and stomatal closure.
  
  
  CONCLUSIONS
  Necrotic lesion formation triggered by ipt activation closely resembles the hypersensitive response. Cytokinins may thus act as signals and/or mediators in plant defence against pathogen attack.
authors: Department of Molecular Biology and Radiobiology, Mendel University in Brno, Zemědělská 1, CZ-613 00 Brno, Czech Republic.
authors: Jan
authors: J
authors: Novák
authors: Jaroslav
authors: J
authors: Pavlů
authors: Ondřej
authors: O
authors: Novák
authors: Vladimíra
authors: V
authors: Nožková-Hlaváčková
authors: Martina
authors: M
authors: Špundová
authors: Jan
authors: J
authors: Hlavinka
authors: Šárka
authors: Š
authors: Koukalová
authors: Jan
authors: J
authors: Skalák
authors: Martin
authors: M
authors: Černý
authors: Břetislav
authors: B
authors: Brzobohatý
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000235:genetics; D016923:Cell Death; D002734:Chlorophyll / Q000378:metabolism; D002736:Chloroplasts / Q000235:genetics / Q000378:metabolism; D003583:Cytokinins / Q000235:genetics / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D020868:Gene Silencing; D054884:Host-Pathogen Interactions; D006861:Hydrogen Peroxide / Q000378:metabolism; D015227:Lipid Peroxidation; D009336:Necrosis / Q000235:genetics; D018384:Oxidative Stress / Q000235:genetics; D010788:Photosynthesis / Q000235:genetics; D010937:Plant Growth Regulators / Q000235:genetics / Q000378:metabolism; D018515:Plant Leaves / Q000166:cytology / Q000235:genetics / Q000502:physiology*; D054046:Plant Stomata / Q000502:physiology; D030821:Plants, Genetically Modified; D014026:Nicotiana / Q000235:genetics / Q000382:microbiology / Q000502:physiology*
chemical_list: D003583:Cytokinins; D010937:Plant Growth Regulators; D002734:Chlorophyll; D003907:Dexamethasone; D006861:Hydrogen Peroxide; D019883:Alkyl and Aryl Transferases; C024274:adenylate isopentenyltransferase
keywords: Cytokinin; Nicotiana tabacum; abscisic acid; hydrogen peroxide; hypersensitive response; jasmonic acid; lipid peroxidation; non-photochemical quenching; pathogenesis-related proteins; photosynthesis; salicylic acid; stomatal conductance
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