Abstract
Suppression subtractive hybridization (SSH) was performed to isolate cDNAs representing genes that are differentially expressed in leaves of Fagus sylvatica upon ozone exposure. 1248 expressed sequence tags (ESTs) were obtained from 2 subtractive libraries containing early and late ozone-responsive genes. Sequences of 1139 clones (91 %) matched the EBI/NCBI database entries. For 578 clones, no putative function could be assigned. Most abundant transcripts were O-methyltransferases , representing 7 % of all sequenced clones. ESTs were organized into 12 functional categories according to the MIPS database. Among them, 12 % (early)/15 % (late) were associated with disease and defence, 19/11 % with cell structure, 4/10 % with signal transduction, and 9/6 % with transcription. The expression pattern of selected ESTs (ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit [rbcS], WRKY-type transcription factor, ultraviolet-B-repressible protein, aquaporine, glutathione S-transferase, catalase, caffeic acid O-methyltransferase, and pathogenesis-related protein 1 [PR1]) was analysed by quantitative real-time RT‐PCR (qRT‐PCR) which confirmed changed transcript levels upon ozone treatment of European beech saplings. The ESTs characterized will contribute to a better understanding of forest tree genomics and also to a comparison of ozone-responsive genes in woody and herbaceous plants.
Key words
Fagus sylvatica
- cDNA - ozone - suppression subtractive hybridization
References
1
Asiegbu F. O., Nahalkova J., Li G..
Pathogen-inducible cDNAs from the interaction of the root rot fungus Heterobasidion annosum with Scots pine (Pinus sylvestris L.).
Plant Science.
(2004);
168
365-372
2
Bhalerao R., Keskitalo J., Sterky F., Erlandsson R., Björkbacka H., Birve S. J., Karlsson J., Gardeström P., Gustafsson P., Lundeberg J., Jansson S..
Gene expression in autumn leaves.
Plant Physiology.
(2003);
131
430-442
3
Chiron H., Drouet A., Claudot A.-C., Eckerskorn C., Trost M., Heller W., Ernst D., Sandermann H..
Molecular cloning and functional expression of a stress-induced multifunctional O -methyltransferase with pinosylvin methyltransferase activity from Scots pine (Pinus sylvestris L.).
Plant Molecular Biology.
(2000);
44
733-745
4
Diatchenko L., Lau Y. F. C., Campell A. P., Chenchik A., Moqadam F., Huang B., Lukyanov S., Lukyanov K., Gurskaya N., Sverdlov E. D., Siebert P. D..
Suppression subtractive hybridization: a method for generating differentially regulated or tissue-specific cDNA probes and libraries.
Proceedings of the National Academy of Sciences of the USA.
(1996);
93
6025-6030
5 Ernst D., Grimmig B., Heidenreich B., Schubert R., Sandermann H.. Ozone-induced genes: mechanisms and biotechnological applications. Smallwood, M. F., Calvert, C. M., and Bowles, D., eds. Plant Responses to Environmental Stress. Oxford; BIOS Scientific Publishers (1999): 33-41
7 Ernst D., Aarts M..
Cis elements and transcription factors regulating gene promoters in response to environmental stress. Sandermann, H., ed. Molecular Ecotoxicology of Plants. Berlin, Heidelberg; Springer-Verlag (2004): 151-176
8
Glick R. E., Schlagnhaufer C. D., Arteca R. N., Pell E. J..
Ozone-induced ethylene emission accelerates the loss of ribulose-1, 5-bisphosphate carboxylase/oxygenase and nuclear-encoded mRNAs in senescing potato leaves.
Plant Physiology.
(1995);
109
891-898
9
Grams T. E. E., Kozovits A. R., Reiter M., Winkler J. B., Sommerkorn M., Blaschke H., Häberle K.-H., Matyssek R..
Quantifying competitiveness in woody plants.
Plant Biology.
(2002);
4
153-158
10
Heidenreich B., Mayer K., Sandermann H., Ernst D..
Mercury-induced genes in Arabidopsis thaliana : identification of induced genes upon long-term mercuric ion exposure.
Plant, Cell and Environment.
(2001);
24
1227-1234
11
Huang X., von Rad U., Durner J..
Nitric oxide induces transcriptional activation of the nitric oxide-tolerant alternative oxidase in Arabidopsis suspension cultures.
Planta.
(2002);
215
914-923
12
Kangasjärvi J., Talvinen J., Utriainen M., Karjalainen R..
Plant defence systems induced by ozone.
Plant, Cell and Environment.
(1994);
17
783-794
13 Kärenlampi L., Skärby L.. Critical levels for ozone in Europe: testing and finalizing the concepts. UN‐ECE workshop report, University of Kuopio, Kuopio, Finland. (1996)
14
Kiefer E., Heller W., Ernst D..
A simple and efficient protocol for isolation of functional RNA from plant tissue rich in secondary metabolites.
Plant Molecular Biology Reporter.
(2000);
18
33-39
15
Kley D., Kleinman M., Sandermann H., Krupa S..
Photochemical oxidants: state of the science.
Environmental Pollution.
(1999);
100
19-42
16
Koch J. R., Scherzer A. J., Eshita S. M., Davis K. R..
Ozone sensitivity in hybrid poplar is correlated with a lack of defense-gene activation.
Plant Physiology.
(1998);
118
1243-1252
17 Langebartels C., Heller W., Ernst D., Lütz C., Payer H.-D., Sandermann H.. Ozone responses of trees: results from controlled chamber exposures at the GSF phytotron. Sandermann, H., Wellburn, A. R., and Heath, R. L., eds. Forest Decline and Ozone: A Comparison of Controlled Chamber and Field Experiments , Ecol. Studies No. 127,. Berlin; Springer (1997): 163-200
18 Langebartels C., Schraudner M., Heller W., Ernst D., Sandermann H.. Oxidative stress and defense reactions in plants exposed to air pollutants and UV‐B radiation. Inzé, D. and Van Montagu, M., eds. Oxidative Stress in Plants. London, New York; Taylor & Francis (2002): 105-135
19
Ludwikow A., Gallois P., Sadowski J..
Ozone-induced oxidative stress response in Arabidopsis : transcription profiling by microarray approach.
Cellular and Molecular Biology Letters.
(2004);
9
829-842
20
Mahalingam R., Gomez-Buitrago A., Eckardt N., Shah N., Guevara-Garcia A., Day P., Raina R., Fedoroff N. V..
Characterizing the stress/defense transcriptome of Arabidopsis .
Genome Biology.
(2003);
4
R20
21
Matyssek R., Sandermann H..
Impact of ozone on trees: an ecophysiological perspective.
Progress in Botany.
(2003);
64
349-404
22
Matyssek R., Wieser G., Nunn A. J., Kozovits A. R., Reiter I. M., Heerdt C., Winkler J. B., Baumgarten M., Häberle K.-H., Grams T. E. E., Werner H., Fabian P., Havranek W. M..
Comparison between AOT40 and ozone uptake in forest trees of different species, age and site conditions.
Atmospheric Environment.
(2004);
38
2271-2281
23
Nunn A. J., Anegg S., Betz G., Simons S., Kalisch G., Seidlitz H., Grams T. E. E., Häberle K.-H., Matyssek R., Bahnweg G., Sandermann H., Langebartels C..
Role of ethylene in the regulation of cell death and leaf loss in ozone-exposed European beech.
Plant, Cell and Environment.
(2005);
28
886-897
24
Pell E. J., Schlagnhaufer C. D., Arteca R. N..
Ozone-induced oxidative stress: mechanisms of action and reaction.
Physiologia Plantarum.
(1997);
100
264-273
25
Pellinen R., Palva T., Kangasjärvi J..
Subcellular localization of ozone-induced hydrogen peroxide production in birch (Betula pendula) leaf cells.
The Plant Journal.
(1999);
20
349-356
26
Pfaffl M. W., Horgan G. W., Dempfle L..
Relative expression software tool (REST©) for group-wise comparison and statistical analysis of relative expression results in real-time PCR.
Nucleic Acids Research.
(2002);
30
e36
27
Plessl M., Rigola D., Hassinen V., Aarts M. G. M., Schat H., Ernst D..
Transcription profiling of the metal-hpyeraccumulator Thlaspi caerulescens (J. & C. PRESL).
Biosciences.
(2005);
60
216-223
28
Rao M. V., Davis K. R..
Ozone-induced cell death occurs via two distinct mechanisms in Arabidopsis : the role of salicylic acid.
The Plant Journal.
(1999);
17
603-614
29
Rishi A. S., Munir S., Kapur V., Nelson N. D., Goyal A..
Identification and analysis of safener-inducible expressed sequence tags in Populus using a cDNA microarray.
Planta.
(2004);
220
296-306
30
Sahr T., Voigt G., Paretzke H. G., Schramel P., Ernst D..
Caesium-affected gene expression in Arabidopsis thaliana.
.
New Phytologist.
(2005);
165
747-754
31 Sandermann H., Wellburn A. R., Heath R. L.. Forest Decline and Ozone. Berlin, Heidelberg, New York; Springer-Verlag (1997)
32
Sandermann H., Ernst D., Heller W., Langebartels C..
Ozone: an abiotic elicitor of plant defence reactions.
Trends in Plant Science.
(1998);
3
47-50
33 Sandermann H., Matyssek R.. Scaling up from molecular to ecological processes. Sandermann, H., ed. Molecular Ecotoxicology of Plants. Berlin, Heidelberg; Springer-Verlag (2004): 207-226
34
Sävenstrand H., Brosché M., Ängehagen M., Strid A..
Molecular markers for ozone stress isolated by suppression subtractive hybridization: specificity of gene expression and identification of a novel stress-regulated gene.
Plant, Cell and Environment.
(2000);
23
689-700
35
Sävenstrand H., Brosché M., Strid A..
Regulation of gene expression by low levels of ultraviolet-B radiation in Pisum sativum : Isolation of novel genes by suppression subtractive hybridisation.
Plant and Cell Physiology.
(2002);
43
402-410
36
Schneiderbauer A., Back E., Sandermann H., Ernst D..
Ozone induction of extensin mRNA in Scots pine, Norway spruce and European beech.
New Phytolologist.
(1995);
130
225-230
37
Schraudner M., Langebartels C., Sandermann H..
Changes in the biochemical status of plant cells induced by the environmental pollutant ozone.
Physiologia Plantarum.
(1997);
100
274-280
38 Schütt P., Schuck H. J., Stimm B.. Lexikon der Forstbotanik. Landsberg; Ecomed (1992)
39
Sharma Y. K., Davis K. R..
The effects of ozone on antioxidant responses in plants.
Free Radical Biology and Medicine.
(1997);
23
480-488
40 Simons S.. Biochemische Effekte und Symptomentwicklung bei Buchen (Fagus sylvatica L.) und Nadelgehölzen unter realen und proportional erhöhten Ozonkonzentrationen. PhD thesis, Ludwig-Maximilians-Universität München. (1993)
41
Tamaoki M., Nakajima N., Kubo A., Aono M., Matsuyama T., Saji H..
Transcriptome analysis of O3 -exposed Arabidopsis reveals that multiple signal pathways act mutually antagonistically to induce gene expression.
Plant Molecular Biology.
(2003);
53
443-456
1 * Both authors contributed equally to this work
D. Ernst
GSF - National Research Center for Environment and Health Institute of Biochemical Plant Pathology
Ingolstädter Landstraße 1
85764 Neuherberg
Germany
Email: ernst@gsf.de
Guest Editor: R. Matyssek