<i>S5H/DMR6</i> Encodes a Salicylic Acid 5-Hydroxylase That Fine-Tunes Salicylic Acid Homeostasis

Zhang, Yanjun; Zhao, Li; Zhao, Jiangzhe; Li, Yujia; Wang, Jinbin; Guo, Ruifeng; Gan, Susheng; Liu, Chang‐Jun; Zhang, Kewei

doi:10.1104/pp.17.00695

Cited by 199 publications

(287 citation statements)

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“…To resolve the SA biosynthetic pathway in plants, we first took a genetic approach. Two Arabidopsis 2-oxoglutarate/Fe (II)dependent dioxygenases (TAIR: AT4G10500 and AT5G24530) were recently characterized as the SA 3-hydroxylase (S3H) and SA 5-hydroxylase (S5H), respectively (Zhang et al, 2013(Zhang et al, , 2017. The single null mutants of S3H and S5H in Arabidopsis, namely s3h and dmr6, exhibit various growth phenotypes and enhanced resistance to pathogens, which are further exacerbated in the s3h/dmr6 double mutant (van Damme et al, 2008;Zhang et al, 2013Zhang et al, , 2017Zeilmaker et al, 2015) ( Figure 1).…”

Section: Resultsmentioning

confidence: 99%

“…Two Arabidopsis 2-oxoglutarate/Fe (II)dependent dioxygenases (TAIR: AT4G10500 and AT5G24530) were recently characterized as the SA 3-hydroxylase (S3H) and SA 5-hydroxylase (S5H), respectively (Zhang et al, 2013(Zhang et al, , 2017. The single null mutants of S3H and S5H in Arabidopsis, namely s3h and dmr6, exhibit various growth phenotypes and enhanced resistance to pathogens, which are further exacerbated in the s3h/dmr6 double mutant (van Damme et al, 2008;Zhang et al, 2013Zhang et al, , 2017Zeilmaker et al, 2015) ( Figure 1). Whereas the s3h plants accumulate SA at a level similar to that in wild-type plants, dmr6 and particularly the s3h/ dmr6 double mutant contain significantly higher levels of free SA, SA 2-O-b-D-glucoside (SAG), and SA glucose ester (SGE) (Zhang et al, 2013(Zhang et al, , 2017 (Figure 2A, Supplemental Figure 2, and Supplemental Data 2).…”

Section: Resultsmentioning

confidence: 99%

“…In this study, we investigate the biochemical functions of PBS3 and EPS1 through untargeted metabolomic characterization of various Arabidopsis SA-metabolism mutants (Zeilmaker et al, 2015;Zhang et al, 2017). Analysis of differentially accumulated metabolites in these mutant lines identified putative intermediates and products of SA metabolism that suggest possible catalytic roles of PBS3 and EPS1.…”

Section: Introductionmentioning

confidence: 99%

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PBS3 and EPS1 Complete Salicylic Acid Biosynthesis from Isochorismate in Arabidopsis

et al. 2019

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Salicylic acid (SA) is an important phytohormone mediating both local and systemic defense responses in plants. Despite over half a century of research, how plants biosynthesize SA remains unresolved. In Arabidopsis, a major part of SA is derived from isochorismate, a key intermediate produced by the isochorismate synthase, which is reminiscent of SA biosynthesis in bacteria. Whereas bacteria employ an isochorismate pyruvate lyase (IPL) that catalyzes the turnover of isochorismate to pyruvate and SA, plants do not contain an IPL ortholog and generate SA from isochorismate through an unknown mechanism. Combining genetic and biochemical approaches, we delineated the SA biosynthetic pathway downstream of isochorismate in Arabidopsis. We found that PBS3, a GH3 acyl adenylase-family enzyme important for SA accumulation, catalyzes ATP-and Mg 2+ -dependent conjugation of L-glutamate primarily to the 8-carboxyl of isochorismate and yields the key SA biosynthetic intermediate, isochorismoyl-glutamate A. Moreover, we discovered that EPS1, a BAHD acyltransferase-family protein with a previously implicated role in SA accumulation upon pathogen attack, harbors a noncanonical active site and an unprecedented isochorismoyl-glutamate A pyruvoylglutamate lyase activity that produces SA from the isochorismoyl-glutamate A substrate. Together, PBS3 and EPS1 form a two-step metabolic pathway to produce SA from isochorismate in Arabidopsis, which is distinct from how SA is biosynthesized in bacteria. This study closes a major knowledge gap in plant SA metabolism and would help develop new strategies for engineering disease resistance in crop plants.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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PBS3 and EPS1 Complete Salicylic Acid Biosynthesis from Isochorismate in Arabidopsis

et al. 2019

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“…Notably, promoters of several genes induced by pathogens and involved in SA metabolism emerged as TPR1 bound (Figure 3). These genes include Isochorismate Synthase 1 ( ICS1 ) and avrPphB Susceptible 3 ( PBS3 ) important for isochorismate derived SA accumulation (Wildermuth et al, 2001; Rekhter et al, 2019; Torrens-Spence et al, 2019), as well as Downy Mildew Resistant 6 ( DMR6 ) encoding an SA deactivating 5-hydroxylase (Zhang et al, 2017). These observations suggest that TPR1 is directly involved in host transcriptional reprogramming of immune responses.…”

Section: Resultsmentioning

confidence: 99%

Genome-wide chromatin binding of transcriptional corepressor Topless-related 1 inArabidopsis

Griebel

Lapin

Kracher

et al. 2020

Preprint

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1 Timely and specific regulation of gene expression is critical for plant responses to 2 environmental and developmental cues. Transcriptional coregulators have emerged 3 as important factors in gene expression control, although they lack DNA-binding 4 domains and the mechanisms by which they are recruited to and function at the 5 chromatin are poorly understood. Plant Topless-related 1 (TPR1), belonging to a 6 family of transcriptional corepressors found across eukaryotes, contributes to 7 immunity signaling in Arabidopsis thaliana and wild tobacco. We performed 8 chromatin immunoprecipitation and sequencing (ChIP-seq) on an Arabidopsis TPR1-9 GFP expressing transgenic line to characterize genome-wide TPR1-chromatin 10 associations. The analysis revealed ~1400 genes bound by TPR1, with the majority 11 of binding sites located at gene upstream regions. Among the TPR1 bound genes, 12 we find not only regulators of immunity but also genes controlling growth and 13 development. To support further analysis of TPR1-chromatin complexes and other 14 transcriptional corepressors in plants, we provide two ways to access the processed 15 ChIP-seq data and enable their broader use by the research community.16 17 Plant materials and growth conditions 121 Stable transgenic lines pTPR1:TPR1-GFP Col-0 (TPR-GFP) and pTPR1:TPR1-HA 122 #3 Col-0 (TPR1-HA) were described previously (Zhu et al., 2010). Plants were grown 123 on soil with a daily 10 h light period (150-200 µE/m 2 s) at 22 °C and 60 % relative 124 humidity. Plant leaf material was harvested after 6 weeks of growth. 125 126 Chromatin immunoprecipitation assays and sequencing (ChIP-seq) 127 ChIP was performed on 2 g leaf tissues of 6-week-old plants according to Gendrel et 128 al. (2005) with modifications (Birkenbihl et al., 2012). Leaf samples were cross-linked 129 twice by infiltration with 1% formaldehyde under a vacuum for 2x 7.5 minutes. Nuclei 130 were disrupted by sonication 10 times for 30 s each and 30 s of cooling in a 131 Bioruptor (Diagenode). IPs were performed using α-GFP (Ab6556, Abcam) 132 antibodies and Protein A agarose beads (Sigma). After elution of immune 133 complexes, cross-linking was reversed with an overnight incubation at 65°C in 134 200 mM NaCl followed by proteinase K digestion. DNA was extracted with phenol-135 chloroform and precipitated with ethanol in the presence of 0.3 M sodium acetate 136 (pH=5.2) and 10 µg/mL glycogen (overnight at -20°C). After centrifugation, DNA 137 pellets were washed with 70% ethanol, air-dried and resuspended in water before 138 PCR. MiniElute PCR Purification Kit (Qiagen, Germany) was used for cleaning ChIP 139 DNA and two rounds of in vitro transcription by T7 RNA polymerase followed 140 according to a linear DNA amplification (LinDA) protocol (Shankaranarayanan et al.

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“…Given the predicted complementarity of AtDMR6 and AtDLO in salicylic acid catabolism (K. Zhang et al , 2013; Y. J. Zhang et al , 2017), particular interest in these results is given by the occurrence of impacting elected mutations in each one of the four scouted genes.…”

Section: Resultsmentioning

confidence: 99%

Mining downy mildew susceptibility genes: a diversity study in grapevine

Pirrello

Zeilmaker²,

Bianco

et al. 2020

Preprint

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AcknowledgmentsCP acknowledges funding from a FEM PhD fellowship at University of Udine, including a support from SciENZA Biotechnologies bv. Conflict of Interest StatementThe authors declare no conflict of interest. AbstractSeveral pathogens continuously threaten viticulture worldwide. Until now, the investigation on resistance loci has been the main trend to understand the interaction between grapevine and mildew causal agents. Dominantly inherited gene-based resistance has shown to be race-specific in some cases, to confer partial immunity and to be potentially overcome within a few years since its introgression. Recently, on the footprint of research conducted on Arabidopsis, the putative hortologues of genes associated with downy mildew susceptibility in this species, have been discovered also in the grapevine genome. In this work, we deep-resequenced four putative susceptibility genes in 190 highly genetically diverse grapevine genotypes to discover new sources of broad-spectrum recessively inherited resistance. The scouted genes are VvDMR6-1, VvDMR6-2, VvDLO1, VvDLO2 and predicted to be involved in susceptibility to downy mildew. From all identified mutations, 56% were Single Nucleotide Polymorphisms (SNPs) in heterozygosity, while the remaining 44% were homozygous. Regarding the identified mutations with putative impact on gene function, we observed ~4% genotypes mutated in VvDMR6-1 and ~8% mutated in VvDMR6-2, only a handful of genotypes that were mutated in both genes. ~2% and ~7% genotypes showed mutations in VvDLO1 and VvDLO2 respectively, and again a few genotypes resulted mutated in both genes. In particular, 80% of impacting mutations were heterozygous while 20% were homozygous.The current results will inform grapevine genetics and corroborate genomic-assisted breeding programs for resistance to biotic stresses.A survey on the genetic diversity of downy mildew susceptibility genes in grapevine varieties and wild species reveals a potential valuable for genomic-assisted breeding as well as tailored gene editing to induce disease resistance.

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S5H/DMR6 Encodes a Salicylic Acid 5-Hydroxylase That Fine-Tunes Salicylic Acid Homeostasis

Cited by 199 publications

References 51 publications

PBS3 and EPS1 Complete Salicylic Acid Biosynthesis from Isochorismate in Arabidopsis

PBS3 and EPS1 Complete Salicylic Acid Biosynthesis from Isochorismate in Arabidopsis

Genome-wide chromatin binding of transcriptional corepressor Topless-related 1 inArabidopsis

Mining downy mildew susceptibility genes: a diversity study in grapevine

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