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ABSTRACT: RNA-Seq analysis of programmed cell death in Arabidopsis thaliana cell suspension culture

PROVIDER: PRJNA926379 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR23190600_1.fastq.gz	Fastqsanger.gz
	SRR23190600_2.fastq.gz	Fastqsanger.gz
	SRR23190601_1.fastq.gz	Fastqsanger.gz
	SRR23190601_2.fastq.gz	Fastqsanger.gz
	SRR23190602_1.fastq.gz	Fastqsanger.gz

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Project description:RNA-Seq Analysis of Arabidopsis thaliana cell suspension cultures undergoing programmed cell death induced by Rose Bengal

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Transcriptional profiling after inhibition of cellulose synthesis by TA and IXB in Arabidopsis thaliana suspension cells

Project description:Transcriptional profiling after inhibition of cellulose synthesis by thaxtomin A and isoxaben in Arabidopsis thaliana suspension cells; Perturbations in the cellulose content of the plant cell wall lead to global modifications in cellular homeostasis, as seen in cellulose synthase mutants or after inhibiting cellulose synthesis. In particular, application of inhibitors of cellulose synthesis such as thaxtomin A (TA) and isoxaben (IXB) initiates a programmed cell death (PCD) in Arabidopsis thaliana suspension cells that is dependent on de novo gene transcription. To further understand how TA and IXB activate PCD, a whole genome microarray analysis was performed on mRNA isolated from Arabidopsis suspension cells exposed to TA and IXB. More than 75% of the genes upregulated by TA were also upregulated by IXB, including genes encoding cell wall-related and calcium-binding proteins, defence/stress-related transcription factors, signalling components and cell death-related proteins. Comparisons with published transcriptional analyses revealed an important subset of genes generally induced in response to various biotic and abiotic stress. Experiment Overall Design: TA, IXB and methanol (control) were added to Arabidopsis thaliana suspension cells three days after sub-culture. Cells were harvested for RNA isolation and frozen in liquid nitrogen after 6 hours of contact with the inhibitors. Samples consisted of four replicates for each condition. A total of 12 Affymetrix GeneChipsÂ® were used in this study, which correspond to 12 RNA samples from the four biological replicates for each of the TA, IXB or methanol addition.

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Project description:Transcriptional profiling after inhibition of cellulose synthesis by thaxtomin A and isoxaben in Arabidopsis thaliana suspension cells Perturbations in the cellulose content of the plant cell wall lead to global modifications in cellular homeostasis, as seen in cellulose synthase mutants or after inhibiting cellulose synthesis. In particular, application of inhibitors of cellulose synthesis such as thaxtomin A (TA) and isoxaben (IXB) initiates a programmed cell death (PCD) in Arabidopsis thaliana suspension cells that is dependent on de novo gene transcription. To further understand how TA and IXB activate PCD, a whole genome microarray analysis was performed on mRNA isolated from Arabidopsis suspension cells exposed to TA and IXB. More than 75% of the genes upregulated by TA were also upregulated by IXB, including genes encoding cell wall-related and calcium-binding proteins, defence/stress-related transcription factors, signalling components and cell death-related proteins. Comparisons with published transcriptional analyses revealed an important subset of genes generally induced in response to various biotic and abiotic stress.

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Project description:We have investigated the genomic response of Arabidopsis cell suspension culture under high light. Our main goal has been twofold: first, to establish whether chloroplasts in Arabidopsis cell suspension culture are functional and, as such, can act as sensors of adverse external stimuli leading to the activation of genomic defence responses in a manner similar to that described in whole plants exposed to a wide range of environmental stresses and; second, to distinguish which of the ROS that would be probably generated in the chloroplasts is predominant. Our functional genomic analysis has led us to conclude that singlet oxygen is the major ROS in Arabidopsis cell suspension culture under high light stress and that singlet oxygen production is responsible for a genomic activation associated with the biosynthesis and signalling pathway of several phytohormones that ultimately triggers accelerated cell death.

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Project description:Despite the critical role of programmed cell death (PCD) in plant development and defence responses, its regulation is not fully understood. It has been proposed that mitochondria may control the early stages of plant PCD, but the details of this regulation are currently unknown. Here, we used Arabidopsis thaliana cell suspension culture, a model system that enables induction and precise monitoring of PCD rates, as well as chemical manipulation of this process to generate a quantitative profile of the alterations in mitochondrial and cytosolic proteomes associated with early stages of plant PCD induced by heat stress. The cells were subjected to PCD-inducing heat levels (10 min, 54oC), with/without the calcium channel inhibitor and PCD blocker LaCl3. The stress treatment was followed by separation of cytosolic and mitochondrial fractions and mass spectrometry-based proteome analysis. Heat stress induced rapid and extensive changes in protein abundance in both fractions and 113 mitochondrial proteins were detected in the cytosol upon PCD induction. Western blot analysis confirmed the release of mitochondrial heat shock protein 60 family members occurring in response to PCD-inducing, but not sublethal level of heat stress, that may indicate their cytosolic role as positive regulators of PCD in plants. In our system, LaCl3 appeared to act downstream of cell death initiation signal, as it did not affect the release of mitochondrial proteins, but instead partially inhibited changes occurring in the cytosolic fraction, including upregulation of proteins with hydrolytic activity. Collectively, these generated data provide new insights into the regulation of cell death and survival decisions in plant cells.

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