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Transgenic strategies for improvement of drought tolerance of cereals to reduce the consequences of water limitations ca

Transgenic strategies for improvement of drought tolerance of cereals to reduce the consequences of water limitations caused by climate change. János Györgyey, Gábor V. Horváth, Dénes Dudits Institute of Plant Biology, Biological Research Centre, Hungarian Academy of Sciences, .

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Transgenic strategies for improvement of drought tolerance of cereals to reduce the consequences of water limitations ca

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  1. Transgenic strategies for improvement of drought tolerance of cereals to reduce the consequences of water limitations caused by climate change János Györgyey, Gábor V. Horváth, Dénes Dudits Institute of Plant Biology, Biological Research Centre, Hungarian Academy of Sciences, Buchanan, Gruissem, Jones: Biochemistry & Molecular Biology of Plants; chapter 22, and results of the Cell cycle and Stress Adaptation Group

  2. N.Sreenivasuluetal./Gene388(2007)1–13

  3. Umezawa et al 2006, 17:113-122 Strategiesforthegeneticengineering ofdroughttolerance.

  4. Buchanan, Gruissem, Jones: Biochemistry & Molecular Biology of Plants; Fig 22.3

  5. Series of responses to drought stress ABA peak Stoma closure Reduced photosynthetic activity Block of cell division, elongation Activation of protective („stress”) genes (DRE, ABRE elements) e.g. ALR Accumulation of osmolytes Long term adaptation

  6. Fig.1 Aschematicrepresentationofcellularsignaltransduction • Pathwaysbetweenstresssignalperceptionandgeneexpressionandthe cis-and trans-elementsinvolvedinstressresponsivegeneexpression.DREB1/CBFandDREB2distinguishtwodifferentsignaltransductionpathwaysinresponsetocoldanddroughtstresses,respectively. DRE:droughtresponsiveelement,ABRE:abscisicacidresponsivebindingelement,MYBRS:MYBrecognitionsite,MYCRS:MYCrecognitionsite,bZIP:basic-domainleucine-zipper • Agarwal et al. PlantCellRep(2006)25:1263–1274

  7. Osmotic stress of wheat plantlets in hydroponics 0 day (10 days old plantlets) Untreated PEG - treated: 100 mOsm 2. days 200 mOsm 4. days 400 mOsm 7. days 9. days 11. days 14. days Sampling

  8. Rice chip – app. 16 000 unigene • hybridised with PEG-treated/ untreated Kobomugi root • samples (day 9) • color flip repeat • app. 5300 spots gave measurable data in both case • >2x induction: more than 1100 spots • >5x induction: 345 spots • >2x repression: more than 400 spots • >5x repression: 77 spots

  9. 6 5 Unknown ACC synthase 5 4 4 3 3 2 2 1 1 0 0 0 day 4 days 9 days 0 day 4 days 9 days 14 5 ADH GST 12 4 10 3 8 6 2 4 1 2 0 0 0 day 4 days 9 days 0 day 4 days 9 days Kobomugi Kobo PEG Plainsmann Pla PEG Relative transcript level of four selected genes exhibiting induction during osmotic stress Q-PCR approved

  10. EXPERIMENTAL SYSTEM FOR EXPOSURE OF WHEAT PLANTLETS TO LONG TERM DROUGHT STRESS IN EXPANDED PERLITE „0 day” (16 day old plantlets) Normalirrigation Reduced irrigation (30%) 1. week 2. week 3. week 4. week Sampling

  11. KOBOMUGI AND PLAISMANN GENOTYPES DIFFER IN VARIETY OF PHYSIOLOGICAL PARAMETERS UNDER WATER STRESS (50% WATER SUPPLY) Kobomugi Plainsmann É. Sárvári et al.

  12. Kobomugi Plainsman V Growth rate of two genotypes under water limitation (30% water supply)

  13. P5CS mRNA in shoots

  14. Transcript level changes in wheat roots during drought adaptation measured on barley macroarray (percentage of 10 500 clones)

  15. 6% 19% 29% 6% 11% 9% 34% 12% Stress and defense Kobomugi Protein synthesis Functional classification of genes upregulated in one genotype only Protein degradation Gene expression Signal transd. Transport Cytosceleton and cell wall Cell growth and division Metabolism Plainsman

  16. Cluster analysis Kobomugi Plainsman Kobomugi Plainsman • putative cyclin-dependent kinase B1-1 • expansin EXPB2 • calmodulin-binding heat-shock protein • xyloglucan endotransglycosylase • Xet3 protein • caffeic acid O-methyltransferase • putative cellulose synthase catalytic subunit • betaine aldehyde dehydrogenase 16

  17. Kobomugi Plainsman Kobomugi Plainsman Kobomugi Plainsman Kobomugi Plainsman 18

  18. Conclusions Divergent drought adaptation strategies of the two genotypes are refelected in their transcript profiles. Long term adaptation is dependent on moderate changes in the expression of large set of genes in a coordinated manner. Transient gene activation is characteristic to Kobomugi, while genes of the more adaptive Plainsmann genoptype exhibit prolonged upregulation. Based on the yield performance and photosynthetic activity, Kobomugi represents escaper strategy while Plainsmann cultivar is capable to maintain physiological functions in harmony with gene expression reprogramming.

  19. Osmotic and Drought Adaptation induced clone Promoter elements in rice orthologue of ODA1 gene Protein function is not known, similar to LEA family and WSI18

  20. Relative transcript levels in roots of Kobomugi during acute drought stress (desiccation)

  21. Standard system for water limitation • Soil → sand:perlite=2:1 • Control plants → 70-80% soil water saturation • Moderately stressed plants → 30-40% soil water saturation • Watering → daily, weight measurment

  22. Relative transcript levels in shoots of Kobomugi after two weeks of moderate drought stress (limited water supply)

  23. Daily change in transcript profile during water limitation in roots of rice • - cv. Sandora • control (100%), stressed (20%) water capacity • 3-3 samples (at 8, 14, 18) • 22 k rice oligo-chip Azsuka Bioryza H Sandora Marilla

  24. Genes induced during the day in rice roots under water limitation And 7 genes with unknown function

  25. Genes repressed during the day in rice roots under water limitation

  26. Genetic engineering of the glyoxalase pathway in tobacco leads to enhanced salinity tolerance Singla-Pareek et al. PNAS 100, 14672-14677(2003)

  27. About the aldose reductase superfamily in general: • Wide range of substrate specificity • Highly conserved structure (NADH or NADPH binding region, catalytic tetrad) • Occurrence: from bacteria to Homo sapiens polyol pathway detoxification of reactive aldehydes

  28. Effects of MsALR overproduction on tobacco plants: • protection against lipid peroxidation under chemical and drought stresses (Oberschall et al. 2000) • protection during drought and UV-B stresses (Hideg et al. 2003) • transgenic plants showed higher tolerance to low temperature and cadmium stress (Hegedűs et al. 2004) • increased tolerance to the effects of high temperature and high light intensity (Horváth and Hideg, unpublished)

  29. Regeneration of the ALR transformants and growth of mature plants Development of first shoots on AAR medium Transgenic plantlets in soil Fertile ALR spikes

  30. IMPROVED PHOTOSYNTHETIC FUNCTION OF WHEAT ALR TRANSFORMANTS (4310-B) AFTER 15 DAYS OF WATER STRESS 120 120 control 100 4310-b 100 80 80 60 Electrontransport (a. u.) Electrontransport (a. u.) 60 40 control 40 4310-b 20 20 0 0 0 500 1000 1500 2000 0 500 1000 1500 2000 PAR (mmol m-2 s-1) PAR (mmol m-2 s-1) E. Hideg et al.

  31. TRANSGENIC WHEAT PLANTS IN THE GREENHOUSE J. Pauket al.

  32. HARVEST INDEX (% DROUGHT STR./UNSTR.) 110 105 THOUSAND KERNEL WEIGHT (% DROUGHT STR./UNSTR.) ALR ACTIVITY IN LEAF EXTRACTS 100 (% DROUGHT STR./UNSTR.) 50 125 40 115 TR288 CTR TR304 105 ALR ACT. (A. U.) 30 (% DROUGHT STR./UNSTR.) 20 95 95 90 85 10 TR288 TR288 CTR CTR TR304 TR304 MsALR EXPRESSING TRANSGENIC WHEAT LINES WITH IMPROVED DROUGHT TOLERANCE

  33. http://www.brc.hu Thank you for your attention

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