Arabidopsis lines with modified ascorbate concentrations reveal a link between ascorbate and auxin biosynthesis
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ABSTRACT:
PROVIDER: PRJNA1261701 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR33522632_1.fastq.gz | Fastqsanger.gz | |||
| SRR33522632_2.fastq.gz | Fastqsanger.gz | |||
| SRR33522633_1.fastq.gz | Fastqsanger.gz | |||
| SRR33522633_2.fastq.gz | Fastqsanger.gz | |||
| SRR33522634_1.fastq.gz | Fastqsanger.gz |
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Project description:Ascorbate is the most abundant water-soluble antioxidant in plants, and it is an essential molecule for normal plant development. It is present in all green plants, with very different concentrations in different plant species. While ascorbate accumulation is a trait of nutritional, and therefore, agronomical interest, the impact of different concentrations over cellular homeostasis remains elusive. In order to shed light over this question, we leveraged Arabidopsis lines with very low ascorbate (vtc2 mutant with 20% of WT ascorbate levels), and low ascorbate concentration (vtc4 mutant with 65% of WT levels), and we generated a line that accumulates 165% of WT levels (vtc2/OE-VTC2). An 80% reduction of ascorbate increased the expression of genes implicated in defense against pathogens, but repressed genes associated with abiotic stress responses. Unexpectedly, lines with increased (165% of WT) and decreased (65% of WT) ascorbate levels shared 85% of induced transcription factors and the GO terms associated with their transcriptional programs. Among the group of genes whose expression is positively correlated with ascorbate content, we identified TAA1/WEI8, a gene encoding a tryptophan aminotransferase that catalyzes the first step of auxin biosynthesis. Using a combination of genetic and pharmacological approaches in fluorescent and histochemical reporter lines for auxin biosynthesis and signaling activity, we revealed that TAA1- and TAA1-RELATED2 (TAR2)-mediated auxin biosynthesis is necessary for plants to cope with increased ascorbate concentration in a light-dependent manner, revealing a new layer of complexity in the regulatory landscape of redox homeostasis.
2025-05-17 | GSE296827 | GEO
Project description:Ascorbate (vitamin C) is an important antioxidant in leaves with multiple functions in photosynthesis and its levels are tightly regulated by light quality. The majority of work to date has focussed on regulation of ascorbate levels via biosynthesis. However, the pathway of ascorbate degradation, which is localised in the apoplast may additionally play a significant role. The degradative pathway begins with ascorbate oxidation by the enzyme ascorbate oxidase (AO) and hence we have examined the impact of manipulation of ascorbate oxidase activity on the content of ascorbate and the acclimation of photosynthesis to changing light levels. Leaf ascorbate content was highly light dependent in Nicotiana tobaccum being double in high compared with low light conditions. In contrast ascorbate oxidase activity was not altered by changing light levels. Transgenic tobacco lines with reduced or enhanced AO activity exhibited no morphological phenotype and photosynthesis under high light was similarly impaired irrespective of the measured AO activity. However further investigation between AO activity and leaf acclimation to changing light intensity revealed a suite of metabolic and transcriptome changes indicating a role for the apoplastic ascorbate pool in chloroplast to nucleus communication. In all genotypes, transcripts associated with ascorbate synthesis and recycling were less abundant following transition from high to low light. However transcripts associated with glutathione recycling, light harvesting, reaction centre function and the Calvin cycle were all increased in abundance. Following the transition from high to low light increases in leaf threonate, an ascorbate degradation product, were proportional to AO activity. A number of chloroplast synthesised amino acids were significantly increased both during high light and following transfer back to LL for 12 h. Similarly, several primary carbohydrates and TCA intermediates were significantly enhanced following high light treatment. In addition to light effects, AO activity had a significant impact on ï¢-alanine, aspartate and threonine. Intriguingly, a large number of fatty acids were reduced in antisense AO and wild-type plants immediately after high light stress however the content of these compounds were significantly increased in overexpressing sense lines. We conclude that AO mediated turnover plays an important role in adjusting cellular ascorbate content to photosynthetic need in a fluctuating light environment and that light mediated signals rapidly adjust that the apoplastic ascorbate pool.
2017-03-01 | E-MTAB-4816 | biostudies-arrayexpress
Project description:We found that auxin stimulates gene expression of DWF4, which encodes a rate-dertermining step in brassinosteroid biosynthesis pathways. This increased gene expressioin subsequently led to elevation of the biosynthetic flux in Arabidopsis roots. To determine the list of genes that are regulated by auxin-synthesizing brassinosteroids, we challenged Arabidopsis seedlings with either auxin only or auxin plus brassinosteroid biosynthetic inhibitor brassinazole. Keywords: Hormone treatment
2011-02-15 | GSE12964 | GEO
Project description:The l-galactose (Smirnoff-Wheeler) pathway represents the major route to l-ascorbic acid (vitamin C) biosynthesis in plants. Arabidopsis thaliana VTC2 and its paralogue VTC5 function as GDP-l-galactose phosphorylases converting GDP-l-galactose to l-galactose-1-P, thus catalyzing the first committed step in the biosynthesis of l-ascorbate. Here we report that the l-galactose pathway of ascorbate biosynthesis described in higher plants is conserved in green algae. The Chlamydomonas reinhardtii genome encodes all the enzymes required for vitamin C biosynthesis via the Smirnoff-Wheeler pathway. We have characterized recombinant C. reinhardtii VTC2 as an active GDP-l-galactose phosphorylase. C. reinhardtii cells exposed to oxidative stress show increased VTC2 mRNA and l-ascorbate levels. We have also shown that enzymatic components of the ascorbate-glutathione system (e.g. ascorbate peroxidase, Mn superoxide dismutase, dehydroascorbate reductase) are up-regulated in response to increased oxidative stress. These results indicate that C. reinhardtii VTC2, like its plant homologs, is a key enzyme in ascorbate biosynthesis in green algae and together with components of the ascorbate recycling system represents the major route in providing protective levels of ascorbate in oxidatively stressed algal cells. Our results suggest that C. reinhardtii cells exposed to oxidative stress conditions produce more ascorbate both by de novo synthesis (Smirnoff-Wheeler pathway) and by recycling via the ascorbate-glutathione cycle.
2012-05-03 | GSE34826 | GEO
Project description:The l-galactose (Smirnoff-Wheeler) pathway represents the major route to l-ascorbic acid (vitamin C) biosynthesis in plants. Arabidopsis thaliana VTC2 and its paralogue VTC5 function as GDP-l-galactose phosphorylases converting GDP-l-galactose to l-galactose-1-P, thus catalyzing the first committed step in the biosynthesis of l-ascorbate. Here we report that the l-galactose pathway of ascorbate biosynthesis described in higher plants is conserved in green algae. The Chlamydomonas reinhardtii genome encodes all the enzymes required for vitamin C biosynthesis via the Smirnoff-Wheeler pathway. We have characterized recombinant C. reinhardtii VTC2 as an active GDP-l-galactose phosphorylase. C. reinhardtii cells exposed to oxidative stress show increased VTC2 mRNA and l-ascorbate levels. We have also shown that enzymatic components of the ascorbate-glutathione system (e.g. ascorbate peroxidase, Mn superoxide dismutase, dehydroascorbate reductase) are up-regulated in response to increased oxidative stress. These results indicate that C. reinhardtii VTC2, like its plant homologs, is a key enzyme in ascorbate biosynthesis in green algae and together with components of the ascorbate recycling system represents the major route in providing protective levels of ascorbate in oxidatively stressed algal cells. Our results suggest that C. reinhardtii cells exposed to oxidative stress conditions produce more ascorbate both by de novo synthesis (Smirnoff-Wheeler pathway) and by recycling via the ascorbate-glutathione cycle. Sampling of Chlamydomonas 2137 exposed to hydrogen peroxide
2012-05-02 | E-GEOD-34826 | biostudies-arrayexpress
Project description:We found that auxin stimulates gene expression of DWF4, which encodes a rate-dertermining step in brassinosteroid biosynthesis pathways. This increased gene expressioin subsequently led to elevation of the biosynthetic flux in Arabidopsis roots. To determine the list of genes that are regulated by auxin-synthesizing brassinosteroids, we challenged Arabidopsis seedlings with either auxin only or auxin plus brassinosteroid biosynthetic inhibitor brassinazole. Keywords: Hormone treatment Arabidopsis seedlings (Columbia ecotype) were grown for 10 d on 1× MS agar-solidified media under long-day conditions (16:8, white light and dark cycle). The seedlings were then transferred to 2 different liquid media containing either 10–7 M 2,4-D or 10–7 M 2,4-D plus 10–6 M brassinazole. After 8 h of treatment, the seedlings were blotted with paper towels to remove excess media and subject to total RNA isolation. Total RNAs isolated from each batch were prepared from 3 replicate seedlings using an RNeasy plant mini kit (Qiagen, Germany).
2011-02-15 | E-GEOD-12964 | biostudies-arrayexpress
Project description:We explore where transcriptional regulation of ascorbate concentration lies in plants. Is it in biosynthesis, recycling, regulation or consumption? Arabidopsis thaliana plants were grown under controlled environment at four photon flux density levels (PFD). Rosettes from plants were harvested at the four PFD levels and over a diurnal cycle and after a step change in PFD and analysed for ascorbate concentration and transcript levels measured by RNAseq. Ascorbate concentrations and expression of genes in the L-galactose ascorbate biosynthesis, recycling, consumption pathways and regulation are presented to provide a full analysis of the control of ascorbate by environmentally modulated gene expression. Ascorbate concentration responded to PFD levels but not to time of day and showed only a small response to change of PFD after 2 days. Of the L-galactose pathway genes, only GDP galactose phosphorylase (GGP) showed a significant response in to different PFDs, time of day and to change in PFD. Other genes also showed limited responses. This study compares gene expression of a range of ascorbate related genes to changes in environment in a unified way and supports the concept that GGP is the key regulatory gene in ascorbate biosynthesis and that post transcriptional regulation is also important.
2017-02-17 | GSE94995 | GEO
Project description:Plant cells exhibit remarkable plasticity of their differentiation states, enabling regeneration of whole plants from differentiated somatic cells. How they revert cell fate and express pluripotency, however, remains unclear. In this study we demonstrate that transcriptional activation of auxin biosynthesis is crucial for reprogramming differentiated Arabidopsis leaf cells. We demonstrate that interfering with the activity of histone acetyltransferases dramatically reduces callus formation from leaf mesophyll protoplasts. Histone acetylation permits the transcriptional activation of PLETHORAs (PLTs), leading to the induction of their downstream target gene YUCCA1 (YUC1) encoding an enzyme for auxin biosynthesis. Auxin biosynthesis is in turn required to accomplish initial cell division through the activation of G2/M phase genes mediated by MYB DOMAIN PROTEIN 3-RELATED (MYB3Rs). We further show that the AUXIN RESPONSE FACTOR 7 (ARF7)/ARF19 and INDOLE-3-ACETIC ACID INDUCIBLE 3 (IAA3)/IAA18-mediated auxin signaling pathway is responsible for cell cycle reactivation by transcriptionally upregulating MYB3R4. These findings provide a mechanistic model of how differentiated plant cells revert their fate and reinitiate the cell cycle to become pluripotent.
2022-07-07 | GSE204767 | GEO
Project description:Ascorbate activates CD30 expression and causes widespread specific demethylation of the epigenome of serum free cultured hESC. The genetic and epigenetic integrity of human Embryonic Stem cells (hESCs) is critical to their future applications in research and medicine. hESC cultured in serum free media can accumulate point mutations, aneuploidy and progressive epigenetic changes over prolonged culture in vitro. We have identified ascorbate as the only molecule in the very widely used Knock-out serum replacement medium that is sufficient to induce expression of CD30, a biomarker for aneuploidy in hESCs. In fact, we show that hESC cultured in the presence of ascorbate for 20 passages not only display demethylation of the CD30 locus, but exhibit widespread and remarkably specific and consistent demethylation of 1,847 genes in both HES2 and HES3 cells. The specific ascorbate induced demethylation changes in the hESC epigenome, of which 86% are shared between the two lines, identify a subset of genes in hESC, including CD30, that are sensitive to serum free culture medium induced epigenetic changes. Experiment. HES2 cells were maintained on 20% Fetal calf serum on mouse embryonic fibroblast feeder layers with mechanical dissection. Using HES2 after 99 mechanical passages (P99), hESC were grown for 17-20 (+17 - +20) passages in the presence of 20% Knock-Out Serum Replacement (Invitrogen) either with (+ASC) or without (-ASC) Ascorbate with enzymatic culturing.
2010-06-10 | GSE16590 | GEO
Project description:Plant cells exhibit remarkable plasticity of their differentiation states, enabling regeneration of whole plants from differentiated somatic cells. How they revert cell fate and express pluripotency, however, remains unclear. Here we show that transcriptional activation of auxin biosynthesis is crucial for reprogramming differentiated leaf cells in Arabidopsis. We demonstrate that intervention of histone acetyltransferases causes severe defects in callus formation from leaf mesophyll protoplasts. Our data suggest that histone acetylation affects transcription of auxin biosynthesis genes. Auxin biosynthesis is in turn required to accomplish initial cell division through the activation of G2/M phase genes mediated by MYB DOMAIN PROTEIN 3-RELATED (MYB3Rs). We further show that AUXIN RESPONSE FACTOR 7 (ARF7)/ARF19 and INDOLE-3-ACETIC ACID INDUCIBLE 3 (IAA3)/IAA18-mediated auxin signaling pathway is responsible for the cell cycle reactivation in protoplasts. These findings provide novel mechanistic insights into how differentiated plant cells revert their fate and reinitiate the cell cycle to exert pluripotency.
2021-10-03 | GSE178645 | GEO