Towards an understanding of the molecular mechanisms of durable and non-durable resistance to stripe rust

1,259 views
1,128 views

Published on

Xianming Chen, USDA-ARS

Published in: Technology, Education
0 Comments
0 Likes
Statistics
Notes
  • Be the first to comment

  • Be the first to like this

No Downloads
Views
Total views
1,259
On SlideShare
0
From Embeds
0
Number of Embeds
100
Actions
Shares
0
Downloads
39
Comments
0
Likes
0
Embeds 0
No embeds

No notes for slide

Towards an understanding of the molecular mechanisms of durable and non-durable resistance to stripe rust

  1. 1. Towards an understanding of the molecular mechanisms of durable and non-durable resistance to stripe rust Xianming ChenUSDA-ARS, Wheat Genetics, Quality, Physiology, and Disease Research Unit, Pullman, WA andDepartment of Plant Pathology, Washington State University
  2. 2. Stripe rust damage can be HUGE Average 2010 3 860 10 20 30 40 50 60 70 80 90 100
  3. 3. Distribution of Stripe Rust in the US in 2010Grain Yield Loss in the US2003: 89 M. Bu (2.42 M. MT)2005: 73 M. Bu (1.99 M. MT)2010: 87 M. Bu (2.38 M. MT)
  4. 4. Damage can be prevented
  5. 5. Yield Loss by Stripe Rust and Increase by Fungicide on Winter Wheat Cultivars, Pullman, WA 2010 Area in 2010 Stripe rust AUDPC Yield (BU/A) YL (%) YI (%) TW (Lb/Bu)Cultivar Acres % Check Fung. Check Fung. Dif. by rust by fung. Check Fung.PS 279 0 0.00 2937.50 1300.75 *** 29.88 65.19 35.31 *** 54.17 118.19 55.44 58.08 **Declo 10,900 0.62 2752.50 566.88 *** 42.15 75.27 33.12 *** 44.00 78.58 54.56 56.67 *Lambert 13,150 0.75 2373.75 409.50 *** 64.43 96.65 32.22 ** 33.34 50.01 56.56 58.04 *Xerpha 159,766 9.13 1624.38 495.75 ** 70.49 95.44 24.95 *** 26.15 35.40 57.83 58.43Tubbs 06 16,200 0.93 2036.25 487.00 *** 78.75 102.32 23.57 *** 23.03 29.93 55.65 56.32Eddy 34,000 1.94 1915.75 330.13 *** 71.37 92.53 21.16 ** 22.87 29.65 60.69 61.92 **Bauermeister 29,500 1.69 1053.75 392.50 *** 67.28 84.75 17.47 * 20.61 25.96 57.97 58.43Paladin 13,000 0.74 2103.13 713.00 *** 63.72 80.03 16.31 * 20.38 25.60 58.29 59.56 **ORCF-103 19,900 1.14 1342.50 450.63 *** 83.07 97.35 14.28 ** 14.67 17.19 56.81 56.78Masami 23,000 1.31 630.75 285.50 *** 86.10 100.64 14.53 ** 14.44 16.88 56.21 56.81Stephens 27,100 1.55 1202.50 196.25 *** 102.07 116.44 14.37 ** 12.34 14.08 58.15 58.61ORCF-102 169,000 9.66 580.88 184.63 ** 90.04 102.57 12.53 * 12.22 13.92 57.48 58.19Brundage 96 39,400 2.25 397.50 82.63 ** 98.37 110.98 12.61 ** 11.36 12.82 57.83 58.61 **Cashup 15,800 0.90 1117.63 291.25 ** 78.78 88.11 9.33 ** 10.59 11.84 59.17 59.14Eltan 309,850 17.71 728.25 265.25 * 85.99 96.05 10.07 10.48 11.71 56.60 56.74Westbred 528 108,100 6.18 513.63 135.00 * 97.44 108.21 10.77 ** 9.95 11.05 59.35 59.95Chuckar 17,800 1.02 60.50 27.00 106.62 114.81 8.19 * 7.14 7.68 59.21 59.56Bruehl 168,900 9.65 78.75 36.00 *** 95.27 102.05 6.78 6.65 7.12 57.13 56.78Finley 42,500 2.43 297.25 71.38 * 61.70 64.98 3.28 5.04 5.31 62.17 62.73Farnum 10,900 0.62 67.25 9.00 * 66.43 67.93 1.51 2.22 2.27 59.49 59.77Madsen 95,100 5.43 39.88 9.00 110.12 111.30 1.18 1.06 1.07 59.59 59.59AP700CL 48,000 2.74 171.38 36.00 * 116.76 115.12 -1.64 -1.43 -1.41 58.75 59.10Rod 56,883 3.25 654.50 242.75 ** 104.87 103.19 -1.69 -1.63 -1.61 57.52 57.41Buchanan 25,700 1.47 345.75 230.38 ** 93.32 89.06 -4.25 ** -4.77 -4.56 57.87 58.19Mean 1042.74 302.01 *** 81.88 95.04 13.17 *** 14.79 21.61 57.93 58.56 *** fMean (Excl. PS 279) (83.11 960.36 258.58 *** 84.14 96.34 12.20 *** 13.07(9.36 ) 17.41 58.04 58.58 ***
  6. 6. In Washington State in 2010:•The various levels of resistance including HTAPresistance were estimated to reduce yield loss frompotentially more than 60% to about 9%.• The application of fungicides in more than 60% of winterand spring wheat acreage in Washington State wasestimated to reduce yield loss further to about 3% (about4.5 million bushels) in average.• The total resistance in all wheat cultivars collectively wasable to save 73 million bushes ($512 million) and thefungicide application further saved 13.7 million bushels($96 million) at the cost of estimated over $27 million.
  7. 7. All-Stage (Seedling) Resistance:Can be detected in seedling stageEffective in all growth stages Yr5 AVSHigh-level resistanceEasy to incorporate into cultivarsNot durable if a single gene is involved
  8. 8. Dr. Orville A. Vogel was the first to develop wheat cultivars with partial resistance to stripe rust IT and % in 2008 Pullman Mt. Vernon Cultivar Release Flow. S. elong Head. Brevor 1949 3 30 5 30 2 20 Omar 1955 8 90 8 60 8 100 Gaines 1960 5 50 8 60 5 60 Nugaines 1965 5 40 8 40 3 40 Luke 1970 2 10 3 15 3 5Photo source: http://cahnrsnews.wsu.edu/reportertools/news/2007/vogel-building-2007-09.html
  9. 9. Dr. Roland F. Linecharacterized high-temperatureadult-plant (HTAP) resistance asthe type of resistance expressedat high post-inoculationtemperatures and at adult-plantstage.
  10. 10. High-Temperature Adult-Plant Resistance Expresses when weather is warm and plants grow old Low to high-level resistance Durable Conferred by quantitative trait loci Relatively difficult to detect and difficult to incorporate into cultivars May not be adequate
  11. 11. Yearrelease Wheat cultivars and their possible source of HTAP resistance1949 Brevor Nord1960 Gaines Desprez1965 Nugaines1970 Luke1971 Hyslop1972 Sprague1976 Raeder McDermid Daws1977 Stephens1979 Walladay1982 Lewjain1983 Hill 811984 Basin John1985 Malcolm Dusty Batum1986 Oveson1987 Frontana Spillman Wakanz1988 Madsen1990 Eltan Kmor1991 Express Bonneville1992 Macvicar Rod1993 Rohde1994 Lambert Alpowa Wawawai1997 Boundary1998 Weatherford Coda Hiller2000 Edwin Cappelle Desprez Hubbard2001 Bruehl Brundage 96 Chukar Finch Gary2002 Tubbs2004 Masami2005 MDM Bauermeister Otis Louise2006 Darwin2007 Xerpha
  12. 12. Why is race-specific all-stage resistancenot durable and nonrace-specific HTAP resistance durable?
  13. 13. Wheat GeneChip 55,052 probe sets Derived from the public content of the T. aestivum UniGene Build #38 Probe sets consist of pairs of 11 perfect match (PM) and mismatch (MM) 25-mer oligonucleotides
  14. 14. Yr5 Experimental Design Near isogenic lines (Isolines) Yr5 AvS  Yr5 (AvSYr5NIL: resistant)  yr5 (AvS: susceptible) Mock- and P. s. tritici-inoculation  Seedling stage (~10 days)  PST-100  Time-course sampling (6, 12, 24, 48 h)  3 biological replicates RNA extraction, labeling and hybridization
  15. 15. Yr5 Data Analysis 120Differentially expressed 100 80 transcripts Yr5 Yr5 60 yr5 40 20 yr5 0 6 12 24 48 Time-point (hpi)
  16. 16. Transcript Separation Yr5 yr5 BiotrophicHR-specific interactiontranscripts 61 54 19 specific transcripts Basal defense transcripts
  17. 17. Transcript Annotation• HR-specific transcripts –Defense signaling via protein kinases –Oxidative burst and HR –PR proteins and phenylpropanoids
  18. 18. Yr39 Experimental Design• Yr39 (resistant) and yr39 (susceptible) RILs from an AvS/Alpowa F7 population• Mock- and P. s. tritici-inoculation –Flag leaf stage (~35 days) –Time-course sampling (12, 24, 48 h) –3 biological replicates• RNA extraction, labeling and hybridization
  19. 19. Fungal Biomass 0.15Accumulation of Pst beta-tubulin mRNA transcripts (1/delta Ct) yr39 0.10 Yr39 * * 0.05 0.00 0 6 12 18 24 30 36 42 48 54 60 66 72 78 84 90 96 102 Time (hpi)
  20. 20. Differential Expression In reference to mock-inoculated controls For Yr39 HTAP resistance interaction  215 transcripts significant at 48 hpi  207 induced and 8 repressed For yr39 compatible interaction  Just 1 transcript induced at 48 hpi
  21. 21. HTAP Resistance-Specific Transcripts• All 99 transcripts were induced• 50.5% involved in pathogen defense and signal transduction• 10 R gene-like transcripts including; – Yr10 all-stage stripe rust resistance protein – Cf2/Cf5 disease resistance protein homolog – Barley stem rust resistance protein Rpg1 – Maize NADPH-dependent HC-toxin reductase Hm1 homolog
  22. 22. HTAP Resistance-Specific Transcripts• Other interesting induced transcripts; – Several protein kinase signaling proteins – Transcriptional regulatory transcripts including a WRKY5 homolog – Five beta-1,3-glucanase transcripts – Wheat pathogen-induced WIR1A protein – Six phenylalanine ammonia-lyase (PAL) transcripts• Quantitative RT-PCR confirmed results
  23. 23. Genes Regulated by Yr5 Race-Specific and Yr39 Nonrace-Specific Resistance to Stripe RustRace-specific Basal Nonrace-specific Basalresistance defense resistance defense yr39 Yr5 yr5 Yr39 61 19 99 1HR 54specific Biotrophic 91 interaction specific
  24. 24. Comparison of transcripts regulated by the Yr5 race-specific all-stage resistance and the Yr39 nonrace-specific HTAP resistanceRace-specific Nonrace-specific resistance resistance Yr5 Yr39 61 99 14
  25. 25. Functional category Yr5 race-specific Yr39 race non- resistance specific resistance• Defense • Resistance proteins - Yr10 homolog - + - Rpg1 homolog - + - Cf2/Cf9 homolog - + - Hm1 homolog - + • Cell wall - WIR1A protein + + - Proline-rich protein + - • Pathogenesis-related proteins - Beta-1,3-glucanase + + - PR protein 10 + - - Thaumatin-like protein + - - Chitinase + - • Phenylpropanoid - Phenylalanine-ammonia lyase + + - UDP-glucosyl transferase - + - Hydroxyanthranilate hydroxyl cinnamoyl - + transferase
  26. 26. Functional category Yr5 race-specific Yr39 race non- resistance specific resistance• Defense • Oxidative stress - Peroxidase + + • Hypersensitive response - S1/P1 nuclease + - - Chromosome condensation factor + - • Miscellaneous - ABC transporter - + - Putative disease resistance protein - + - Latex protein allergen - +• Signal transduction - Protein kinase + + - Receptor protein kinase - + - Calmodulin protein + +• Transcription - WRKY5 homolog - + - Myb transcription factor + -
  27. 27. Meta-analysis Design• Custom oligonucleotide microarray – 116 significant transcripts from Yr5 results – 207 significant transcripts from Yr39 results• Aim to identify common/unique gene expression signatures involved in each resistance
  28. 28. Resistance Type Comparison (More Genes of Races-Specific vs. Nonrace-specific)• 8 wheat genotypes with race-specific resistance – Yr1, Yr5, Yr7, Yr8, Yr9, Yr10, Yr15 and Yr17• 4 genotypes with nonrace-specific resistance – Yr18, Yr29, Yr36 and Yr39• Mock and incompatible interaction – Seedling and adult plant stage
  29. 29. Race-specific Resistance• Seedling stage phenotype effect – Combined genotype data• 28 transcripts significant – P <0.10 and fold change >2.0• Compared to 116 transcripts in Yr5 response – Meta-analysis narrowed the gene list
  30. 30. Transcript AnnotationFunctional category Putative function Fold change p valueDefense Putative disease resistance protein 2.45 0.017Defense Putative disease resistance protein 2.36 0.017Defense - alkaloid 28 transcripts annotated • 21 of the Reticuline oxidase 2.01 0.078Defense - cell wall Pathogen induced WIR1A protein 2.15 0.000 – 15 (71%) involved in defense/signalingDefense - oxidative stress Blue copper-binding protein 4.11 0.000Defense - oxidative stress Blue copper-binding protein 2.35 0.012Defense - oxidative stress Peroxidase 2.54 0.022Defense - oxidative stress Peroxidase 2.71 0.089Defense - phenylpropanoid Phenylalanine ammonia-lyase 2.13 0.004Defense - phenylpropanoid Phenylalanine ammonia-lyase 5.43 0.001Defense - PR protein Beta-1,3-glucanase 2.04 0.087Defense - PR protein PR protein 10 2.01 0.003Defense - R protein NB-ARC domain containing protein 2.66 0.024Signal transduction Calmodulin-binding protein 2.82 0.055Signal transduction LRR-containing extracellular glycoprotein 2.57 0.001Transcription Transcription factor 2.40 0.000
  31. 31. Nonrace-specific resistance• Only detectable at adult-stage• Zero significant transcripts for nonrace- specific resistance phenotype effect• Directly compared race-specific resistance to nonrace-specific resistance – 5 transcripts significant for race-specific resistance – 1 transcript significant for nonrace-specific resistance
  32. 32. Race-specific resistanceFunctional Category Putative Function Fold change p valueDefense - cell wall Hydroxyproline-rich glycoprotein 10.78 0.000Defense - R protein NB-ARC domain containing protein 2.22 0.006Signal transduction Protein kinase 5.50 0.000Unknown No homology 4.38 0.000Unknown No homology 2.43 0.000 Nonrace-specific resistanceFunctional category Putative function Fold change p valueTransport Nonclathrin coat protein 2.16 0.000
  33. 33. Relationships of Yr genes based on common and unique transcripts in response to stripe rust infection Yr1 Yr5 Yr17 Yr15 Yr8 Yr10 Yr18 Yr39 Yr9 Yr29
  34. 34. Conclusions and Perspectives In comparison with race-specific all-stage resistance, nonrace- specific HTAP resistance is contributed by a relatively great number of defense-related genes, which may explain the durability of HTAP resistance. Genes contributing to all-stage resistance are induced fast and their transcription levels increased dramatically in the infection process, while those contributing to HTAP resistance are induced more slowly and their transcription changes are less dramatic. All-stage resistances mediated by different R genes tend to share many common defense genes, while HTAP resistances-mediated by different genes do not have many defense genes in common. Transcription factors identified in these studies may play key roles in the networks of plant defense. Further characterization of these genes may enhance our understanding of molecular mechanisms of different types of resistance.
  35. 35. Cloning a Pleiotropic Drug Resistance/ ABC Transporter Gene from Alpowa Ta.6990.1.S1_at 1.29* Mean log2 fold change (Pst inoculated – mock 1.4 1.2 inoculated) 1 0.8 * Significant 0.6 0.25 0.27 0.4 0.02 0.2 0 24hai 48hai HTAP Resistance (Yr39) Susceptible (yr39)
  36. 36. Genes with similar sequence to Ta.6990.1.S1 Ta.6990.1.S1_at is likely a PDR [Pleiotropic Drug Resistance]-type ABC [ATP Binding Cassette] transporter
  37. 37. ABC Geneassociated withTa.6990.1.S1_at Yr18/Lr34 Ta.6990.1.S1_at Yr18/Lr34 Size (kb) 7.4 11.8 Introns 18 24 Chromosome 7A* 7D*• Lr34-A, a homolog of Lr34, is located on 7A, but its sequence is dissimilar from the Ta.6990.1 associated gene.The gene associated with the Ta.6990.1.S1 transcript issubstantially different from Yr18/Lr34 (47% similar) eventhough both are PDR- type ABC transporters.
  38. 38. Thank You Tristan Coram Xueling Huang Meinan Wang Andrea Dolezal

×