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Transcriptome analyses of mock- and Pst DC3000-inoculated Arabidopsis thaliana Col-0 plants at normal and elevated temperature

ABSTRACT: We performed RNA sequencing of mock-inoculated and Pseudomonas syringae pv. tomato (Pst) DC3000-infected A. thaliana (Col-0 accession) plants at normal (23C) and elevated (30C) temperatures. 4-week-old Col-0 plants were pre-incubated at 23C and 30C for 48h and then leaves were syringe-infiltrated with either mock (MgCl2) or DC3000 bacterial inoculum. Plants were incubated at their respective temperatures (23C or 30C) for another 24h post-inoculation before tissue collection for RNA extraction. RNA samples for submitted for RNA sequencing and we found different clusters of DC3000-regulated genes that were downregulated, upregulated and unchanged at elevated temperature. Temperature-downregulated DC3000-induced genes were enriched for a whole suite of defense-related genes, including those essential for host salicylic acid (defense hormone) biosynthesis and accumulation.

ORGANISM(S): Arabidopsis thaliana

PROVIDER: GSE152072 | GEO | 2022/05/05

REPOSITORIES: GEO

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Project description:We have implemented an integrated Systems Biology approach to analyze overall transcriptomic reprogramming and systems level defense responses in the model plant Arabidopsis thaliana during an insect (Brevicoryne brassicae) and a bacterial (Pseudomonas syringae pv. tomato strain DC3000) attack. The main aim of this study was to identify the attacker-specific and general defense response signatures in the model plant Arabidopsis thaliana while attacked by phloem feeding aphids or pathogenic bacteria. Defense responses and networks, unique and specific for aphid or Pseudomonas stresses were identified. Our analysis revealed a probable link between biotic stress and microRNAs in Arabidopsis and thus opened up a new direction to conduct large-scale targeted experiments to explore detailed regulatory links among them. The presented results provide a first comprehensive understanding of Arabidopsis - B. brassicae and Arabidopsis - P. syringae interactions at a systems biology level. Arabidopsis thaliana (ecotype Colombia-0) seeds were sown into 6-cm-diameter pots filled with a sterile soil mix (1.0 part soil and 0.5 part horticultural perlite). Plants were kept in growth chambers VM-CM-6tsch VB 1514 (VM-CM-6tch Industrietechnik GmbH, Germany) with a 16/8 h (light/dark) photoperiod at 22/18 M-BM-0C, 40/70% relative humidity, and 70/0 mmol m-2 s-1 light intensity. The Pseudomonas syringae pv. tomato strain DC3000 culture was grown overnight in 10 ml of Kings B solution supplemented with antibiotics rifampicin (50 M-NM-<g mlM-bM-^HM-^R1) and kanamycin (25 M-NM-<g mlM-bM-^HM-^R1). Overnight culture was washed once in 10 mM MgCl2 and final cell densities were adjusted to approximately 0.20 at 600 nm (approximately 1.5 M-CM-^W 108 cfu mlM-bM-^HM-^R1) in 10 mM MgCl2. Plants were mock-challenged with 10 mM MgCl2 or inoculated with DC3000 strain, 3-4 leaves were infiltrated on the abaxial surface with a needleless 1-ml syringe.Whole rosettes were cut at the hypocotyls and harvested from Pseudomonas infested and mock-infected plants after 72 hours treatment. 4 biological replicates were prepared from each treatment, each containing rosettes from 15 individual plants. Differences in transcriptional responses were measured by comparing genes expression of Pseudomonas infected plants against mock-infected control plants.

Project description:Plant 9-lipoxygenases (9-LOX) and α-dioxygenases (α-DOX) initiate the synthesis of oxylipins after bacterial infection. Here, the role of these enzymes in plant’s defense was investigated using individual Arabidopsis thaliana lox1 and dox1 mutants and a double lox1 dox1 mutant. Studies with Pseudomonas syringae pv tomato (Pst) revealed the enhanced susceptibility of lox1 to the virulent strain Pst DC3000 and the partial impairment of lox1 and dox1 mutants to activate systemic acquired resistance. Notably, both defects were enhanced in the lox1 dox1 plants as compared with individual mutants. We found that pre-treatment with 9-LOX- and -DOX-generated oxylipins protected plant tissues against bacterial infection. The strongest effect in this respect was exerted by 9-ketooctadecatrienoic acid (9-KOT), which is produced from linolenic acid by 9-LOX. Quantification of 9-KOT revealed its accumulation after bacterial infection. The levels were reduced in lox1 and lox1 dox1 plants but strongly increased in the dox1 mutant due to metabolic interaction of the two pathways. Transcriptional analyses indicated that 9-KOT pre-treatment modifies hormone homeostasis during bacterial infection. The nature of the changes detected suggested that 9-KOT interfers the hormonal changes caused by bacterial effectors. This notion was substantiated by the finding that 9-KOT failed to reduce the growth of PstDC3000hrpA, a mutant compromised in effector secretion, and of the avirulent strain Pst DC3000 avrRpm1. Further support to the action of the 9-LOX- and -DOX-oxylipin pathways as modulators of hormone homeostasis was the observation that lox1 dox1 seedlings are hypersensitive to the growth-inhibitory effect of ABA and showed enhanced activation of ABA-inducible marker genes. Two experiments, Col-0 treated with 9-KOT vs. Col-0 treated with water and Col-0 treated with 9-KOT vs. Col-0 treated with water after treatment with Pst DC3000. Biological replicates: 4 control replicates treated , 4 9-KOT treated replicates; 4 control treated and Pst DC3000 replicates and 4 9-KOT treated and Pst DC3000 replicates

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Transcriptome analyses of mock- and Pst DC3000-inoculated Arabidopsis thaliana Col-0 plants at normal and elevated temperature

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