rcs5-mediated spot blotch resistance in barley is conferred by wall-associated kinases that resist pathogen manipulation

ABSTRACTPlant biotrophic pathogen disease resistances rely on immunity receptor-mediated programmed cell death (PCD) responses, but specialized necrotrophic/hemi-biotrophic pathogens hijack these mechanisms to colonize the resulting dead tissue in their necrotrophic phase. Thus, immunity receptors can become necrotrophic pathogen dominant susceptibility targets but resistance mechanisms that resist necrotroph manipulation are recessive resistance genes. The barley rcs5 QTL imparts recessive resistance against the disease spot blotch caused by the hemi-biotrophic fungal pathogen Bipolaris sorokiniana. The rcs5 genetic interval was delimited to ~0.23 cM, representing an ~234 kb genomic region containing four wall-associated kinase (WAK) genes, designated HvWak2, Sbs1, Sbs2 (susceptibility to Bipolaris sorokiniana1&2), and HvWak5. Post-transcriptional gene silencing of Sbs1&2 in susceptible barley cultivars resulted in resistance showing dominant susceptibility function. Allele analysis of Sbs1&2 from resistant and susceptible barley cultivars identified sequence polymorphisms associated with phenotypes in their primary coding sequence and promoter regions, suggesting differential transcriptional regulation may contribute to susceptibility. Transcript analysis of Sbs1&2 showed nearly undetectable expression in resistant and susceptible cultivars prior to pathogen challenge; however, upregulation of both genes occurred specifically in susceptible cultivars post-inoculation with a virulent isolate. Apoplastic wash fluids collected from barley infected with a virulent isolate induced Sbs1, suggesting regulation by an apoplastic-secreted effector. Thus, Sbs1&2 function as B. sorokiniana susceptibility targets and non-functional alleles or alleles that resist induction by the pathogen mediate rcs5-recessive resistance. The sbs1&2 alleles underlying the rcs5 QTL that the pathogen is unable to manipulate are the first resistance genes identified against spot blotch.SIGNIFICANCE STATEMENTThe rcs5 locus in barley confers a high level of seedling resistance and a moderate level of adult plant resistance to spot blotch. It is part of a complex that has provided durable spot blotch resistance in many North American barley cultivars (cv) for more than 50 years. Genetic characterization and positional cloning of rcs5 identified the dominant susceptibility genes, Sbs1 and Sbs2 (susceptibility to Bipolaris sorokiniana 1 and 2) as wall-associated kinases. These genes are hijacked by the hemibiotrophic pathogen in its necrotrophic phase to induce programmed cell death, facilitating disease development. We report the first spot blotch resistance/susceptibility genes cloned that function via alleles that cannot be specifically induced and hijacked by virulent isolates of the pathogen.

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Molecular Mapping of Loci Conferring Susceptibility to Spot Blotch and Resistance to Powdery Mildew in Barley Using the Sequencing-Based Genotyping Approach

Phytopathology ◽

10.1094/phyto-08-19-0292-r ◽

2020 ◽

Vol 110 (2) ◽

pp. 440-446 ◽

Cited By ~ 3

Author(s):

Yueqiang Leng ◽

Mingxia Zhao ◽

Jason Fiedler ◽

Antonín Dreiseitl ◽

Shiaoman Chao ◽

...

Keyword(s):

Powdery Mildew ◽

Molecular Mapping ◽

Recombinant Inbred Lines ◽

Major Gene ◽

Bipolaris Sorokiniana ◽

Spot Blotch ◽

Blumeria Graminis ◽

Seedling Stage ◽

Barley Cultivars ◽

Trait Locus

Spot blotch (SB) caused by Bipolaris sorokiniana and powdery mildew (PM) caused by Blumeria graminis f. sp. hordei are two important diseases of barley. To map genetic loci controlling susceptibility and resistance to these diseases, a mapping population consisting of 138 recombinant inbred lines (RILs) was developed from the cross between Bowman and ND5883. A genetic map was constructed for the population with 852 unique single nucleotide polymorphism markers generated by sequencing-based genotyping. Bowman and ND5883 showed distinct infection responses at the seedling stage to two isolates (ND90Pr and ND85F) of Bipolaris sorokiniana and one isolate (Race I) of Blumeria graminis f. sp. hordei. Genetic analysis of the RILs revealed that one major gene (Scs6) controls susceptibility to Bipolaris sorokiniana isolate ND90Pr, and another major gene (Mla8) confers resistance to Blumeria graminis f. sp. hordei isolate Race I, respectively. Scs6 was mapped on chromosome 1H of Bowman, as previously reported. Mla8 was also mapped to the short arm of 1H, which was tightly linked but not allelic to the Rcs6/Scs6 locus. Quantitative trait locus (QTL) analysis identified two QTLs, QSbs-1H-P1 and QSbs-7H-P1, responsible for susceptibility to spot blotch caused by Bipolaris sorokiniana isolate ND85F in ND5883, which are located on chromosome 1H and 7H, respectively. QSbs-7H-P1 was mapped to the same region as Rcs5, whereas QSbs-1H-P1 may represent a novel allele conferring seedling stage susceptibility to isolate ND85F. Identification and molecular mapping of the loci for SB susceptibility and PM resistance will facilitate development of barley cultivars with resistance to the diseases.

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A Compromised Mlo Pathway Affects the Response of Barley to the Necrotrophic Fungus Bipolaris sorokiniana (Teleomorph: Cochliobolus sativus) and Its Toxins

Phytopathology ◽

10.1094/phyto.2001.91.2.127 ◽

2001 ◽

Vol 91 (2) ◽

pp. 127-133 ◽

Cited By ~ 135

Author(s):

Jagdish Kumar ◽

Ralph Hückelhoven ◽

Ulrich Beckhove ◽

Subrahmaniam Nagarajan ◽

Karl-Heinz Kogel

Keyword(s):

Cell Death ◽

Cell Wall ◽

Powdery Mildew ◽

Bipolaris Sorokiniana ◽

Spot Blotch ◽

Cochliobolus Sativus ◽

Fungal Culture ◽

Wild Type ◽

Powdery Mildew Fungus ◽

Resistance Traits

In search of new durable disease resistance traits in barley to control leaf spot blotch disease caused by the necrotrophic fungus Bipolaris sorokiniana (teleomorph: Cochliobolus sativus), we developed macroscopic and microscopic scales to judge spot blotch disease development on barley. Infection of barley was associated with cell wall penetration and accumulation of hydrogen peroxide. The latter appeared to take place in cell wall swellings under fungal penetration attempts as well as during cell death provoked by the necrotrophic pathogen. Additionally, we tested the influence of a compromised Mlo pathway that confers broad resistance against powdery mildew fungus (Blumeria graminis f. sp. hordei). Powdery mildew-resistant genotypes with mutations at the Mlo locus (mlo genotypes) showed a higher sensitivity to infiltration of toxic culture filtrate of Bipolaris sorokiniana as compared with wild-type barley. Mutants defective in Ror, a gene required for mlo-specified powdery mildew resistance, were also more sensitive to Bipolaris sorokiniana toxins than wild-type barley but showed less symptoms than mlo5 parents. Fungal culture filtrates induced an H2O2 burst in all mutants, whereas wild-type (Mlo) barley was less sensitive. The results support the hypothesis that the barley Mlo gene product functions as a suppresser of cell death. Therefore, a compromised Mlo pathway is effective for control of biotrophic powdery mildew fungus but not for necrotrophic Bipolaris sorokiniana. We discuss the problem of finding resistance traits that are effective against both biotrophic and necrotrophic pathogens with emphasis on the role of the anti-oxidative system of plant cells.

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ETUDE DE LA TOLERANCE DE CULTIVARS D’ORGE A LA TACHE HELMINTHOSPORIENNE

Canadian Journal of Plant Science ◽

10.4141/cjps87-018 ◽

1987 ◽

Vol 67 (1) ◽

pp. 153-157 ◽

Cited By ~ 3

Author(s):

D. DOSTALER ◽

G. J. PELLETIER ◽

L. COUTURE

Keyword(s):

Hordeum Vulgare ◽

Grain Yield ◽

Seed Weight ◽

Bipolaris Sorokiniana ◽

Spot Blotch ◽

Grain Weight ◽

Disease Development ◽

Barley Cultivars ◽

Moderate Disease ◽

Key Words Barley

This study was conducted to assess tolerance of barley (Hordeum vulgare) cultivars to spot blotch. The barley cultivars Parkland, Bonanza and Laurier inoculated with spores of Bipolaris sorokiniana developed approximately the same level of spot blotch symptoms in the field. Natural disease development was minimal in plots treated with mancozeb; it was intermediate in uninoculated and untreated plots. Reduction of grain yield and grain weight differed with each cultivar. Disease had the most depressing effects on Parkland. Disease severity was highest on Laurier which suffered the least decrease in grain yield and grain weight compared to the checks. Yield and seed weight of Bonanza were not affected by low or moderate disease levels (checks) but were reduced when symptoms were severe (inoculations).Key words: Barley, cultivars, spot blotch, Bipolaris sorokiniana, tolerance, mancozeb

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Mutations in a barley cytochrome P450 gene enhances pathogen induced programmed cell death and cutin layer instability

PLoS Genetics ◽

10.1371/journal.pgen.1009473 ◽

2021 ◽

Vol 17 (12) ◽

pp. e1009473

Author(s):

Gazala Ameen ◽

Shyam Solanki ◽

Lauren Sager-Bittara ◽

Jonathan Richards ◽

Prabin Tamang ◽

...

Keyword(s):

Cell Death ◽

Cytochrome P450 ◽

Programmed Cell Death ◽

Plant Immunity ◽

Susceptibility Gene ◽

Bipolaris Sorokiniana ◽

Post Inoculation ◽

Cytochrome P450 Gene ◽

P450 Gene ◽

Pigmented Lesions

Disease lesion mimic mutants (DLMMs) are characterized by the spontaneous development of necrotic spots with various phenotypes designated as necrotic (nec) mutants in barley. The nec mutants were traditionally considered to have aberrant regulation of programmed cell death (PCD) pathways, which have roles in plant immunity and development. Most barley nec3 mutants express cream to orange necrotic lesions contrasting them from typical spontaneous DLMMs that develop dark pigmented lesions indicative of serotonin/phenolics deposition. Barley nec3 mutants grown under sterile conditions did not exhibit necrotic phenotypes until inoculated with adapted pathogens, suggesting that they are not typical DLMMs. The F2 progeny of a cross between nec3-γ1 and variety Quest segregated as a single recessive susceptibility gene post-inoculation with Bipolaris sorokiniana, the causal agent of the disease spot blotch. Nec3 was genetically delimited to 0.14 cM representing 16.5 megabases of physical sequence containing 149 annotated high confidence genes. RNAseq and comparative analysis of the wild type and five independent nec3 mutants identified a single candidate cytochrome P450 gene (HORVU.MOREX.r2.6HG0460850) that was validated as nec3 by independent mutations that result in predicted nonfunctional proteins. Histology studies determined that nec3 mutants had an unstable cutin layer that disrupted normal Bipolaris sorokiniana germ tube development.

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Mutations in a barley cytochrome P450 gene enhances pathogen induced programmed cell death and cutin layer instability

10.1101/2021.03.09.434546 ◽

2021 ◽

Author(s):

G. Ameen ◽

S. Solanki ◽

L. Sager-Bittara ◽

J. Richards ◽

P. Tamang ◽

...

Keyword(s):

Cell Death ◽

Cytochrome P450 ◽

Programmed Cell Death ◽

Defense Mechanisms ◽

Bipolaris Sorokiniana ◽

Post Inoculation ◽

Cytochrome P450 Gene ◽

P450 Gene ◽

Lesion Mimic ◽

Pigmented Lesions

ABSTRACTDisease lesion mimic mutants (DLMMs) are characterized by spontaneous development of necrotic spots with various phenotypes designated as necrotic (nec) mutants in barley. The nec mutants were traditionally considered to have aberrant regulation of programmed cell death (PCD) pathways, which have roles in plant immunity and development. Most barley nec3 mutants express cream to orange necrotic lesions contrasting them from typical spontaneous DLMMs that develop dark pigmented lesions indicative of serotonin/phenolics deposition. Also, barley nec3 mutants grown under sterile conditions did not exhibit necrotic phenotypes until inoculated with adapted pathogens suggesting that they are not typical DLMMs. The F2 progeny of a cross between nec3-γ1 and variety Quest segregated as a single recessive gene post inoculation with Bipolaris sorokiniana, the causal agent of the disease spot blotch. Nec3 was genetically delimited to 0.14 cM representing 16.5 megabases of physical sequence containing 149 annotated high confidence genes. RNAseq and comparative analysis of wild type and five independent nec3 mutants identified a single candidate cytochrome P450 gene (HORVU.MOREX.r2.6HG0460850) that was validated as nec3 by independent mutations that result in predicted nonfunctional proteins. Histology studies determined that nec3 mutants had an unstable cutin layer that disrupted normal Bipolaris sorokiniana germ tube development.AUTHOR SUMMARYAt the site of pathogen infection, plant defense mechanisms rely on controlled programmed cell death (PCD) to sequester biotrophic pathogens that require living cells to extract nutrient from the host. However, these defense mechanisms are hijacked by necrotrophic plant pathogens that purposefully induce PCD mechanism to feed from the dead cells facilitating further disease development. Thus, understanding PCD responses is important for resistance to both classes of pathogens. We characterized five independent disease lesion mimic mutants of barley designated necrotic 3 (nec3) that show aberrant regulation of PCD responses upon pathogen challenge. A cytochrome P450 gene was identified as Nec3 encoding a Tryptamine 5-Hydroxylase that functions as a terminal serotonin biosynthetic enzyme in the Tryptophan pathway of plants. The nec3 mutants have disrupted serotonin biosynthesis resulting in expansive PCD, necrotrophic pathogen susceptibility and cutin layer instability. The nec3 mutants lacking serotonin deposition in pathogen induced necrotic lesions show expansive PCD and disease susceptibility suggesting a role of serotonin to sequester PCD and suppress pathogen colonization. The identification of Nec3 will facilitate functional analysis to elucidate the role serotonin plays in the elicitation or suppression of PCD immunity responses to diverse pathogens and effects it has on cutin layer biosynthesis.

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The In-Silico Development of DNA Markers for Breeding of Spring Barley Varieties That Are Resistant to Spot Blotch in Russia

Agriculture ◽

10.3390/agriculture10110505 ◽

2020 ◽

Vol 10 (11) ◽

pp. 505

Author(s):

Irina V. Rozanova ◽

Nina M. Lashina ◽

Vadim M. Efimov ◽

Olga S. Afanasenko ◽

Elena K. Khlestkina

Keyword(s):

Dna Markers ◽

Root Rot ◽

Spring Barley ◽

Significant Snps ◽

Bipolaris Sorokiniana ◽

Spot Blotch ◽

Yield Losses ◽

Barley Cultivars ◽

Northwestern Region ◽

The Impact

The fungal pathogen Cochliobolus sativus Drechs. Ex Dastur, anamorph Bipolaris sorokiniana (Sacc.) Shoemaker is one of the most common barley pathogens worldwide and causes spot blotch and root rot in barley. Spot blotch is considered to be the major biotic stress hampering the commercial production of barley. During high disease severity, which occurs in the northwestern region of Russia once every three to four years, yield losses for barley may reach 40%. An increase in common root rot severity results in yield losses that can reach 80%. The goal of the current study was to identify significant markers that can be employed as diagnostic DNA markers to breed C. sativus pathogen-resistant varieties of barley. In 94 spring barley cultivars and lines, the resistance of seedlings and adult plants to the impact of C. sativus on their leaves and roots was investigated. Five genomic regions associated with resistance to Spot blotch were identified (on chromosome 1H (50–61.2 cM), 2H (68.7–69.68 cM), 3H (18.72–26.18 cM), 7H (7.52–15.44 cM)). No significant loci were determined to be associated with root rot. According to obtained data, 11 significant SNPs were converted into KASP markers and 6 markers located on chromosome 3H were determined to possess good accuracy and the potential to be employed in marker-assisted selection.

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