Uv radiation-and-molecular-effects

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Uv radiation-and-molecular-effects

  1. 1. The Effects of Ultraviolet Radiation and Canopy Shading on Grape Berry Biochemistry & Molecular Biology Professor Brian Jordan Professor of Plant Biotechnology Agriculture and Life Sciences Faculty Lincoln University
  2. 2. Responses of Plants to Light Light Photosynthesis Sugars other organic compounds Information leaf growth stem growth germination, etc. flowering dormancy plant habit, etc. direction of growth Small amounts of light Daily duration of light Direction of light
  3. 3. 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 300 400 500 600 700 800 Wavelength (nm) Spectralirradiance(relativeunits) 900 1000 Plants Red & far redBlue UV-A UV-B
  4. 4. Ultraviolet Penetration through the Stratospheric Ozone Layer UV-A 380-315nm UV-B 315-280nm UV-C <280nm O3layer 0%100% Earth’s surface PAR 700nm – 380nm
  5. 5. Photoperception to gene expression Photoperception Signal Transduction Gene Expression
  6. 6. UV-B Photoreceptor UV-B Specific Photoreceptor Signal Transduction Non-Specific Via ROS Via DNA damage Changes to gene expression
  7. 7. H2O2 PR genes JA O2 - PDF1.2 Ethylene SA Transcription factors Photosynthetic genes H2O2 Chloroplast signal, electron transport/ photophosphorylation UV-B Peroxidase NADPH oxidase Receptor Signal Transduction Pathways ? NO Ca2+ /CaM Phosphorylation NOS Chs
  8. 8. Role of UV/Light in Grape Development and Wine Quality • Effect on “ageing” of white wines in New Zealand • Changes to polyphenolic compounds • Changes to amino acids/protein content • Impact on aroma/flavour (methoxypyrazines) • Lipoxygenase as an example of molecular approach
  9. 9. Vineyard experiments • UVA+, UVB+ screen • UVA+, UVB- screen • UV- screen • No frame • No leaf removal, no frame 0 20 40 60 80 100 250 275 300 325 350 375 400 Wavelength nm %Transmission UV+ UVA+ UV-
  10. 10. UV-B Damage No UV-B Damage
  11. 11. UV-absorbing compounds Total peak area Integratedarea@352nm Total peak areaTotal peak area Integratedarea@352nm
  12. 12. Amino Acid Metabolism and Implications for Wine Industry UV (and PAR) NITROGEN (Uptake and assimilation) AMINO ACIDS Methoxypyrazines: amino acids as precursors to flavour and aroma compounds Phenolics: amino acids as precursors – implicated in ageing and bitterness in white wine Amino acid composition and implications for fermentation bouquet and yeast assimilable nitrogen Glutathione: implicated in the prevention of browning process Valine, isoleucine, leucine Phenylalanine, tyrosine, tryptophan All amino acids except proline Cysteine, glutamate, glycine
  13. 13. Amino Acid Composition Glutamine Proline Arginine Alanine Serine Glutamate Arginine Proline Glutamine Alanine Threonine Serine Increasing Amounts ChardonnayChardonnay SauvignonSauvignon blancblanc
  14. 14. Light regulation of nitrogen metabolism • Light regulates the conversion of glutamate into glutamine in the chloroplast • This involves the GOGAT pathway and requires ATP • This assimilation of nitrogen then provides amino acids/amines to the fruit Glutamate Glutamine
  15. 15. Amino acids Glutamine 0 20 40 60 80 100 120 Lo UV UV-A All UV %ofno-frame
  16. 16. Amino acids Glutamic acid 0 10 20 30 40 50 60 70 80 90 No pluck Lo UV UV-A All UV No frame µM
  17. 17. Major aroma chemicals • 3-mercaptohexanol/3- mercaptohexanal acetate – Tropical fruit and Citrus aromas • Methoxypyrazines – Green/green-pepper or capsicum aromas
  18. 18. Present Understanding: Synthesis of Thiol Precursors Lipids and Fatty Acids in Cell Membranes 5/6 Carbon Backbone eg, s-3- (hexan-1-ol)- Glutathione LOX HPL etc Non Volatile s-cysteine Conjugate Precursor Grape Metabolism through Berry Development and in Response to the Environment Changes during Must Fermentation Release of Aroma Volatiles Primarily by Yeast VERAISON Hard Solid Berry Soft Berry at Harvest ‘Membrane Turnover’ GSTs
  19. 19. COOH OOH 13(S)-HPOT CHO (3Z)-hexenal COOHOHC (9Z)-12-oxododec-9-enoic acid CHO OH COOH OHC COOH OH COOH HOOC OH CHO O(O)H Traumatin (9Z)-12-hydroxy-9-dodecenoic acid Traumatic acid (3Z)-hexen-1-ol (2E)-hexenal (2E)-4-hydro(pero)xy-2-hexenal (2E)-hexen-1-ol HPL IF ADH ADH ADH IF LOX? 9(S)-HPOT COOH HOO HPL COOHOHC 9-oxononanoic acid CHO (3Z,6Z)-nonadienal CHO OH (2E,6Z)-nonadienal (3Z,6Z)-nonadien-1-ol IF ADH HOOC CH3 a-linolenic acid S t o r a g e lip id s B io lo g ic a l m e m b r a n e s F r e e fa tt y a c id s 13-LOX9-LOX 9(S)-HPOT - (10E, 12Z, 15Z)-9-hydro(pero)xy-10,12,15-octadecatrienoic acid; 13(S)-HPOT - (9Z,11E,15Z)-13-hydro(pero)xy-9,11,15-octadecatrienoic acid; HPL - hydroperoxide lyase; LOX - lypoxygenase; ADH - alcohol dehydrogenase; IF - isomerization factor; LOX-HPL pathway
  20. 20. 13-LOXs Type I 9-LOXs Type I Type II13- LOXs LO X1 Gm 1 LO X1 G m 2 LO X1 Ah 1 LO X1 Ps 2 LOX 1 G m 6 LOX1 Gm 7 L OX1 G m 3 LO X1 Ps 3 LO X1 Lc 1 LO X 1 G m 4 LOX1Gm5 LOX1 Cs1 LOX1Cs2 LOX1St2 LOXLVv LOX1At2 LOX1St1 LOX1Le1 LOX1Nt1 LOX1Prd 1 LO X1 A t1 L O X 1 C a 1 LO XM Vv LOX B Vv LO XC Vv L OX 1 Hv 1 LOX 1 Zm 3 LO X1 O s 1 LO X1 Zm 1 LOX 2 Zm 6 LO XD Vv L OX 2 At 2 LOX 2 A t 3 L O X2 St 2 LOXO VvLOXR Vv LO X2 At 4 LO XP V v L O X 2 O s 1 LO X2 Zm 1 LOX2 Hv 1 LOX2Os2 LOX2At1 LOX2Bn2 LOX2St1 LOX2Pod1 LOX2Pod2 LOXJVv LOXK VvLOXA Vv L OX E Vv LOX F Vv LO XG Vv LOX H V v LOXI Vv Phylogenetic analysis of grape LOXs and characterised LOXs from other plants
  21. 21. Proportional distribution of grape LOXs in different berry fractions Relative expression of four berry expressed LOXs SB berry expressed LOXs 0% 20% 40% 60% 80% 100% VvLOXA VvLOXC VvLOXD VvLOXO Proportionaltranscriptabundance Skin Pulp Seed
  22. 22. Relative gene expressions of berry expressed LOXs during development
  23. 23. Relative gene expressions of berry expressed LOXs during upon wounding
  24. 24. I – berries with obvious signs of infection, NI – berries closely located to the infected, Control – healthy berries distantly located from the infected. Relative LOX gene expressions in SB berries infected with Botrytis
  25. 25. Vmax 16.0546 0.6008 Km 2.1092 0.3049 Vmax 7.5836 0.1551 Km 0.8196 0.0981 Vmax 6.6200 Km 0.5582
  26. 26. pH effect on recombinant VvLOXA activity
  27. 27. pH effect on recombinant VvLOXO activity
  28. 28. Methoxypyrazines • Little is known about their biosynthesis – Thought to derive from amino acid biosynthesis • Accumulate up until veraison • Degrade after veraison and with exposure of grape bunches to light • At low concentrations (ng.L-1 ) contribute to green/green- pepper aromas
  29. 29. UV responses & wine quality
  30. 30. +UV No leaf No No No UV removal frame UV-B UV responses & wine quality
  31. 31. Effects of UV and Leaf Removal on Wine Quality • Methoxypyrazine levels low in juice at harvest, but high early in grape development: control of gene expression from amino acid precursors • Amino acid composition different in juice in response to light environment • Regulation of proline biosynthesis important for fermentation • Flavonoids accumulate with UV exposure: role of transcription factors • Lipoxygenase pathway: complex gene family and expression pattern
  32. 32. Acknowledgements Grape Biotechnology and UV Research • Jason Wargent, Lancaster University, UK • Scott Gregan • Stephen Stilwell • Andriy Podolyan (Ph.D.) • Jim Shinkle, Trinity University, USA • Dr Rainer Hofmann • Dr Chris Winefield • Professor Brian Jordan (Programme Leader) Support From: • Foundation for Research, Science & Technology • NZ Royal Society/MoRST COST-ACTION 858 • Marlborough Wine Research Centre, Auckland University & Plant & Food Research • New Zealand Wine Industry • Lincoln University

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