cracking the codes: genetic basis of nonhost resistance of barley to heterologous rust fungi
DESCRIPTION
Rients Niks, Wageningen UniversityTRANSCRIPT
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Cracking the codes: Genetic basis of nonhost resistance of barley to heterologous rust fungi
Rients Niks, Hossein Jafary, Thierry MarcelLaboratory of Plant Breeding, Wageningen University
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P. triticina
Barley
Wheat
Barley is host to Puccinia hordei, but nonhost to leaf rusts of rye, wheat and grasses.
Host status:
Can we make wheat a “quasi-nonhost” to P. triticina ?
What do we know about Nonhost resistance?
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Non-host resistance:
• constitutive• morphological
• physiological
• induced
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cell wall PAMP
PAMP = Pathogen associated molecular pattern
RLKperception of non-self
RLK = Receptor-like kinase, recognizing the PAMP
signal transduction
Unadapted pathogen
defence reaction
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cell wall PAMP
PAMP = Pathogen associated molecular pattern
RLKperception of non-self
RLK = Receptor-like kinase, recognizing the PAMP
signal transduction
Adapted pathogen
defence reaction
effectors
Suppression of defence
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cell wall PAMP
PAMP = Pathogen associated molecular pattern
RLKperception of non-self
RLK = Receptor-like kinase, recognizing the PAMP
signal transduction
Unadapted pathogen
defence reaction No suppression of defence
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Plant cell
Pathogen
The players
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The defencePerception
Signal transduction
Defence
PAMP
PRR
SA, JA
PR proteins,Chitinases, Callose deposition, Silicium deposition
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Switch off the alarm
1 0 6 71 0 6 7Effectors
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Host-status: degree of match between effectors of pathogens and targets in the plant
Which are the targets?
Relevant effectors are presumably transcription factors or specific proteinases
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Barley
P. hordei
host
host
non-host
non-host
P. triticina
Wheat
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Barley
P. hordei P. triticina
host
host
non-host
non-host
++
--
Monogenic or polygenic?
Wheat
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Barley
P. hordei
host
host
non-host
non-host
+ + + + + +
+ + + + + + +- - - - - -
- - - - - - -
Monogenic or polygenic?
Wheat
P. triticina
Are the same genes responsible for resistance to other unadapted rusts?
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Barley
P. hordei
host
host
non-host
non-host
+ + + + + +
+ + + + + + +- - - - - -
- - - - - - -
Wheat
P. triticina
- - - - - - -- - - - - - -- - - - - - -
+ + + + + ++ + + + + ++ + + + + +
Do all barley accessions share the same genes?
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- - - - + + -- - - + - - +Barley
P. hordei
host
host
non-host
non-host
+ + + + + +
+ + + + + + +Wheat
P. triticina
- + - - - - -+ + + - - - -
+ + + + + ++ + + + + ++ + + + + +
Do all barley accessions share the same genes?Are such nonhost genes present also for the adapted pathogen?
- - - - - -
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+ - + + + ++ + + + - +
- + + + + + - - - + - - +Barley
P. hordei
host
host
non-host
non-host
+ + + + + + +Wheat
P. triticina
- - - - + + -- + - - - - -+ + + - - - -
+ + + + + +
Are such nonhost genes present also for the adapted pathogen?
- - - - - -
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- + + + + + - - - + - - +Barley
P. hordei
host
host
non-host
non-host
+ + + + + + +Wheat
P. triticina
- - - - + + -- + - - - - -+ + + - - - -
+ + + + + ++ - + + + ++ + + + - +
- - - - - -
Are such nonhost genes present also for the adapted pathogen? Quantitative basal host resistance?
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Basal resistance (partial resistance)
“what makes the plant less than extremely susceptible”
non-hypersensitive resistance
L94
Vada
Puccinia hordei
Vada
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Barley-Puccinia is an excellent system to study those questions
P. triticina
Barley
Wheat
rare barley accession, L94
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Quantification of host status of barley to rusts of grasses
• 110 barley accessions:
landraces, modern cultivars, Asian, American, African, …; Hordeum spontaneum
• Rust species collected from other Hordeum species, or from Triticum, Agropyron, etc
• Inoculation with 3x as much inoculum than “normal” (= in P. hordei studies)
• All tests in seedling stage
Atienza et al (2004)Atienza et al (2004)
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0%10%20%30%40%50%60%70%80%90%
100%
P. persistens
P. triticina
P. hor.-secalini
P. hordei-bulbosi
P. graminis lolii
P. hor.-murini
P. cor. 5 ff.spp.
P. holci
U. dactylidis
P. bromina
P. recondita
P. vulpiana
P.rec. alopecuri
Blumeria graminis f. sp. tritici
P. hordei (1.2.1)
> 500 pustules
> 100 pustules
11-100 pustules
3-10 pustules
<3 pustules, more flecks
< 3 pustules, few flecks
(Near)-non-host status of barley to Puccinia rustsFull nonhost-statusNear-nonhost-status
Atienza et al (2004)
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Development of hyper-susceptible experimental lines
P. triticina
several barley accessions , like L94
SusPtrit
Atienza et al (2004)
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SusPtritVada
C. Capa
P. tr iticina
P. hordei-bulbosi
P. hordei-murini
P. gram. lolii
P. persistens
susceptibleimmune
immune
P. recondita
P. holci
P. bromi
etc
immuneimmune
immune
Atienza et al 2004
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SusPtritVada C. CapaXX
3 mapping populations x 4 heterologous rusts
Oregon Wolfe barleys
Jafary et al, MPMI 2006Jafary et al, Genetics 2008
Quantitative inheritance! Jafary et al, MPMI 2006Jafary et al, Genetics 2008
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Vada x SusPtrit
Immunity of Vada to each heterologous rust depends on a different set of QTLs!
Jafary et al, MPMI 2006Jafary et al, Genetics 2008
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Hypothesis:
• These QTLs may contain genes (or encode proteins) that are targets to the effectors of rust fungal effectors
• In that case, each rust has different targets to suppress PAMP triggered defence!
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Host range genes to Puccinia
Jafary et al, MPMI 2006Jafary et al, Genetics 2008
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Hypothesis:
• These QTLs may contain genes (or encode proteins) that are targets to the effectors of rust fungal effectors
• In that case, each rust has different targets to suppress PAMP triggered defence!
• What kind of genes are these targets? Are they defencegenes? Same genes as genes for basal resistance to P. hordei?
L94
Vada
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Host range genes to Puccinia
Jafary et al, MPMI 2006Jafary et al, Genetics 2008
BINs of 5 cM eachWhole genome = 220 BINs
Some markers developed on PRX motif
Chi-square test on co-incidence of peak markers of nonhost QTLs with QTLs for partial resistance and PRX genes
Gonzalez et al PLoS-ONE 2010
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30FHDCFV and VSCADI conserved motifs
Possible role for peroxidase genesAminoacid sequences of 119 barley peroxidases http:/ / peroxidase.isb-sib.ch
150 bp
Gonzalez et al PLoS-ONE 2010
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31MseI/FH2Y
MseI/ FH1C
Motif-directed profiling for prx-motifs
• RILs from L94 x Vada (96 RILs)• RILs from SusPtrit x Vada (118 RILs)MseIFH2Y-680L/ 676V
MseIFH2Y-650L/ 640V
MseIFH2Y-678L
MseIFH1C-660S/ 657V
MseIFH1C-650S/ 590V
Gonzalez et al PLoS-ONE 2010
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Prx markers and resistance QTLs placed on integrated linkage map
Prx markers occur in ~ 40 clusters
Significant tendency of resistance QTLs to map in prx-gene clusters
Gonzalez et al PLoS-ONE 2010
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33
Prx QTLph1 QTLbg QTLnh QTLdh QTLdp QTLplh QTLkw QTLtw QTLyi
Marker no. 200 19 23 63 52 15 31 13 18 24
BIN no. 63 18 23 47 39 9 28 11 13 23
O (E) 11 (5.2) 14 (6.7) 22 (13.4)
15 (11.5) 3 (2.6) 12 (8.3) 5 (3.2) 5 (3.8) 9 (6.8)
χ2 9.9 * 12.6** 9.9* 1.8 0.2 2.8 1.4 0.6 1.1
Gonzalez et al PLoS-ONE 2010
Significant association of QTLs for basal resistance with location of PRX genes
PRX not associated with QTLs for agronomic traits
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Barley
P. hordei
host
host
non-host
non-host
- + + + + +
+ + + + + + +- - - - - -
- - - + + - +
Wheat
P. triticina
- - - - + + -+ + - - - + ++ + + + - - -
+ + + + + ++ - + + + ++ + + + - +
• Quantitative inheritance
• Occasionally R-gene for HyR
• In each parental combination the resistance resides on different loci
• For each rust species a different set of genes
• Significant co-incidence with basal resistance to P. hordei
• Significant co-incidence with defence related ESTs, especially PRX
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How to proceed?
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Mapping and fine-mapping QTLs
QTL-NILs developed:
L94 background:
Rphq2Rphq3Rphq4Rphq2 + Rphq3RnhqAll from Vada
SusPtrit background:
Rphq2 (Vada)Rphq3 (Vada)
Rphq11 (Steptoe)Rphq16 (Dom)
Rnhq (L94)Rhnq (Vada)
Yeo et al, unpubl.
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NILs
P. h
orde
i.1.2
.1
P. h
orde
i.Co-
4
P. h
or s
ec. F
P. h
or s
ec W
P. h
or s
ec G
P. g
ram
. Lol
ii
P. h
or m
urin
i
P. p
ersi
sten
s
P. tr
titic
ina.
Fla
min
go
P. tr
titic
ina.
Swiz
P. g
raqm
tri
t
P. h
or b
RLP (3 x) RLP (3 x) RIF (1 x) RIF (1 x) RIF (1 x) RIF (2 x) RIF (2 x) RIF (2 x) RIF (3 x) RIF (1 x) RFF (2 x) RIF (2 x)
SusPtrit 100 100 100 100 100 100 100 100 100 100 100 100
Su-Rphq2 106 100 98 70 81 108 103 140 110 144 102 126
Su-Rphq3 106 101 126 111 85 31 80 114 57 59 91 88
Su-Rphq11 105 103 27 33 26 32 44 54 87 25 66 2
Su-Rphq16 108 106 165 107 87 102 113 189 90 123 88 91
Su-Rnhq.L 100 100 96 53 61 57 93 75 85 131 116 87
Su-Rnhq.v 101 101 54 55 75 54 87 65 88 122 123 71
L94 100 100 100 100 100 100 100 HR 100 100 100 100
L94-Rphq2 106 108 14 123 76 115 72 HR 87 48 115 84
L94-Rphq3 105 106 9 42 36 31 80 HR 35 52 49 86
L94-Rnhq 100 99 2 12 16 3 28 HR 75 23 107 93
Charactirization of QTLs in NILs
LP IF
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Defence genes: have general effectiveness: those effective to P. triticina should also be effective to P. hordei.
However, small sequence variations in coding region or promoter may determine whether effector can reprogramme the gene expression?
What may be the targets of the effectors?
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Barley/ Puccinia offers an excellent opportunity to investigate the specificity aspect of non-host resistance:
- Diploid autogamous plant species, with great diversity
- Extensive knowledge on markers, micro-arrays, dense linkage maps
- Is near-nonhost to several different rust species and forms
- Related to other important cereal crops
Gonzalez et al PLoS-ONE 2010
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Barley-Puccinia rust is effective system to understand the inheritance of host status of plants
Arabidopsis is not (yet) used to tackle this question
We uncover natural variation (no mutagenesis)
TILLING is no immediate option unknown which is the functional allele (susc or res?)
redundancy of QTLs for nonhost resistance
Most common mapping populations do not segregate (but OWB!)
LD mapping on cultivar collection is no option
BAC libraries available • Vada: resistance alleles of 28 QTLs • Cebada Capa: resistance allele of 13 QTLs• SusPtrit: QTL-genes can be susceptibility factors
Concluding remarks
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IPKDr. Rajeev K. Varshney Prof. Dr. Andreas GranerDr. Nil SteinsDr. Marion Röder Dr. Patrick Schweizer
URGV-INRADr. Boulos Chalhoub Cécile Huneau
StudentsJerome DurandThijs van DijkAlice LoriauxFreddy Yeo Kuok San
Wageningen University-Lab of Plant BreedingRichard G.F. VisserAnton VelsReza Aghnoum Ana Maria GonzalezZuzana Kohutova
SCRIDr. Ingo Hein Dr. Robbie Waugh
Acknowledgements