Transcriptomic changes in aquaporin PIP1;2 deficient Arabodposis plants
ABSTRACT: The leaf transcriptome of the Arabidopsis thaliana aquaporin gene PIP1;2 T-DNA insertion line was compared to that of control plants. In total 730 genes were found to be differentially regulated. This regulation pattern was compared to mild drought stress and low CO2 Affymetrix data to elucidate whether loss of the aquaporin resembles transcriptomic changes of drought stress or lack of CO2 supply. Mild drought stress data were obtained from Harb A, Krishnan A, Ambavaram MMR, Pereira A (2010) Molecular and Physiological Analysis of Drought Stress in Arabidopsis Reveals Early Responses Leading to Acclimation in Plant Growth. Plant Physiology 154: 1254-1271 (GSE24177). Low CO2 data were obtained from Oliver E. Bläsing, Yves Gibon, Manuela Günther, Melanie Höhne, Rosa Morcuende, Daniel Osuna, Oliver Thimm, Björn Usadel, Wolf-Rüdiger Scheible, and Mark Stitt (2005) Sugars and Circadian Regulation Make Major Contributions to the Global Regulation of Diurnal Gene Expression in Arabidopsis. The Plant Cell, Vol. 17, 3257-3281 (GSE3423). 2 samples examined: wildtype and atpip1;2-1 mutant
Project description:Genome-wide transcriptional profiling of Arabidopsis thaliana to a combination of heatwave and drought under ambient and elevated CO2. Goal of this study was elucidate the transcriptional responses to a combination of heat wave and drought, and to see how these responses are modifed under future climate (high) CO2. Climate changes increasingly threaten plant growth and productivity. Such changes are complex and involve multiple environmental factors, including rising CO2 levels and climate extreme events. As the molecular and physiological mechanisms underlying plant responses to realistic future climate extreme conditions are still poorly understood, a multiple organizational level-analysis (i.e. eco-physiological, biochemical and transcriptional) was performed, using Arabidopsis exposed to incremental heat wave and water deficit under elevated CO2.The climate extreme resulted in biomass reduction, photosynthesis inhibition, and considerable increases in stress parameters. Photosynthesis was a major target as demonstrated at the physiological and transcriptional levels. In contrast, the climate extreme treatment induced a protective effect on oxidative membrane damage, most likely as a result of strongly increased lipophilic antioxidants and membrane-protecting enzymes. Elevated CO2 significantly mitigated the negative impact of a combined heat and drought, as apparent in biomass reduction, photosynthesis inhibition, chlorophyll fluorescence decline, H2O2 production and protein oxidation. Analysis of enzymatic and molecular antioxidants revealed that the stress-mitigating CO2 effect operates through up-regulation of antioxidant defense metabolism, as well as by reduced photorespiration resulting in lowered oxidative pressure. Therefore, exposure to future climate extreme episodes will negatively impact plant growth and production, but elevated CO2 is likely to mitigate this effect. Transcriptome analysis was performed by Agilent Arabidopsis (V4) 4x44K platform which represented all known genes in the Arabidopsisgenome. Experiments were performed using a modified loop design (Knapen et al., 2009). This design consisted of total 8 arrays; sample from each treatment was labelled once and has 4 biological replicates, two of which were labelled in red and two in green
Project description:Drought is one of the most detrimental environmental stresses to which plants are exposed. Especially mild drought is relevant to agriculture and significantly affects plant growth and development. In plant research, mannitol is often used to mimic drought stress and study the resulting responses. In growing leaf tissue of plants exposed to mannitol-induced stress, a highly-interconnected gene regulatory network is induced. However, early signaling and associated protein phosphorylation events that likely precede part of these transcriptional changes are largely unknown. Here, we performed a full proteome and phosphoproteome analysis on growing leaf tissue of Arabidopsis plants exposed to mild mannitol-induced stress and captured the fast (within the first half hour) events associated with this stress. Based on this in-depth data analysis, 167 and 172 differentially abundant and unique proteins and phosphorylated sites were found back, respectively. Finally, we identified H(+)-ATPASE 2 (AHA2) and CYSTEINE-RICH REPEAT SECRETORY PROTEIN 38 (CRRSP38) as novel regulators of shoot growth under osmotic stress.
Project description:Plant homeodomain (PHD) finger proteins affect growth and development by regulating transcription and reading epigenetic modifications of histones, but their functions in abiotic stress responses remain largely unclear. Here we characterize seven Arabidopsis thaliana Alfin1-like PHD finger proteins (ALs) in the response to abiotic stresses. ALs localize to the nucleus and repress transcription. Except AL6, all the ALs bind to G-box element. Changes of the amino acids at positions 34 and 35 in AL6 cause the loss of G-box binding ability. Expression of the ALs responded differently to osmotic stress, salt, cold and abscisic acid treatments. AL5 was induced by multiple stresses, and AL5-overexpressing plants showed higher tolerance to salt, drought and freezing stress than Col-0. Also, al5 mutants showed reduced stress tolerance. ChIP-Seq assay helps find the direct targets of AL5. Polyclonal antibody of AL5 protein was used to perform ChIP experiment. Two samples were analyzed, AL5 OE sample and its knock out mutant. Data was analyzed as OE sample Vs mutant sample and help find targets of AL5 protein.
Project description:UV radiation is a ubiquitous component of solar radiation that affects plant growth and development. Analysis of natural variation in response to UV radiation revealed significant differences among natural accessions of Arabidopsis thaliana. However, the genetic basis of this is to a large extent unknown. Here, we analyzed the response of Arabidopsis accessions to UV radiation stress by performing RNA-sequencing of three UV sensitive and three UV resistant accessions. The genome-wide transcriptional analysis revealed a large number of genes significantly up- or down-regulated only in sensitive or only in resistant accessions, respectively. Mutant analysis of few selected candidate genes suggested by the RNA-sequencing results indicate a connection between UV radiation stress and plant-pathogen-like defense responses. Examination of transcriptional changes in response to UV treatment in Arabidopsis natural accessions
Project description:With frequent fluctuations in global climate, plants often experience co-occurring dry-wet cycles and pathogen infection and this combination adversely affects plant survival. In the past, some studies indicated that morpho-physiological responses of plants to the combined stress are different from the individual stressed plants. However, interaction of drought stressed or drought recovered plants with pathogen has not been widely studied at molecular level. Such studies are important to understand the defense pathways that operate as part of combined stress tolerance mechanism. In this study, Arabidopsis plants were exposed to individual drought stress (soil drying at 40% FC, D), Pseudomonas syringae pv tomato DC3000 (PStDC3000), infection and their combination. Plants recovered from drought stress were also exposed to PStDC3000. Beside we have also infiltrated P. syringae pv tabaci (PSta, non-host pathogen) individually or in combination with drought stress. Using Affymetrix WT gene 1.0 ST array, global transcriptome profiling of plants leaves under individual drought stress and pathogen infection was compared with their combination. Results implicate that plants exposed to combined drought and pathogen stress experience a new state of stress where each combination of stressor and their timing defines the plant responses and thus should be studied explicitly. Global transcriptional analysis in Arabidopsis leaves exposed to individual and combined drought and pathogen stress. Overall design: Microarray based global gene expression analysis was carried out in Arabidopsis leaves after exposure to individual and combined stress treatments. Arabidopsis plants were exposed to drought stress (D), PStDC3000 infection (P, 1d and 6d), PSta infection (NH, 1d). For treatments involving combined stress, PStDC3000 was infiltrated on drought stressed plants (DP, 1d), plants were exposed to drought stress upon PStDC3000 infiltration (PD, 1d), PSta was infiltrated on drought stressed plants (DNH, 1d), and PStDC3000 was infiltrated on drought recovered plants (DRP). Six plants per treatment were used such that leaf tissue from three plants made one biological replicate. Two biological replicates were hybridized for each treatment.
Project description:RNA sequencing on the third true Arabidopsis leaves of plants exposed to mild drought stress from 12 days after stratification onwards. Samples were harvested at 4AM, 12PM and 8PM on day 14, and at 4AM on day 15.
Project description:Drought is an important environmental factor affecting plant growth and biomass production. Despite this importance, little is known on the molecular mechanisms regulating plant growth under water limiting conditions. The main goal of this work was to investigate, using a combination of growth and molecular profiling techniques, how Arabidopsis thaliana leaves adapt their growth to prolonged mild osmotic stress. Fully proliferating, expanding and mature leaves were harvested from plants grown on plates without (control) or with 25mM mannitol (osmotic stress) and compared to seedlings at stage 1.03. Overall design: Total RNAs were extracted using Trizol method from vegetative part of seedlings at stage 1.03 and leaves that are fully proliferating, expanding or mature. All plants were grown in vitro on medium without (0mM) or with mannitol (25mM) in three independent biological experiments. Each sample was pooled from multiple plants and multiple plates in one experiment. RNA samples were submitted to ATH1 array hybridization.
Project description:Drought is an important environmental factor affecting plant growth and biomass production. Despite this importance, little is known on the molecular mechanisms regulating plant growth under water limiting conditions. The main goal of this work was to investigate, using a combination of growth and molecular profiling techniques, how Arabidopsis thaliana leaves adapt their growth to prolonged mild osmotic stress. Fully proliferating, expanding and mature leaves were harvested from plants grown on plates without (control) or with 25mM mannitol (osmotic stress) and compared to seedlings at stage 1.03. Total RNAs were extracted using Trizol method from vegetative part of seedlings at stage 1.03 and leaves that are fully proliferating, expanding or mature. All plants were grown in vitro on medium without (0mM) or with mannitol (25mM) in three independent biological experiments. Each sample was pooled from multiple plants and multiple plates in one experiment. RNA samples were submitted to ATH1 array hybridization.
Project description:Plants engineered for abiotic stress tolerance may soon be commercialized. The engineering of these plants typically involves the manipulation of complex multigene networks and may therefore have a greater potential to introduce pleiotropic effects than the simple monogenic traits that currently dominate the plant biotechnology market. Drought- tolerant Arabidopsis thaliana were engineered through overexpression of the transcription factor ABF3 in order to investigate unintended pleiotropic effects. In order to eliminate position effects, the Cre/lox recombination system was used to create control plant lines that contain identical T-DNA insertion sites but with the ABF3 transgene excised. This additionally allowed us to determine if Cre recombinase can cause unintended effects that impact the transcriptome. Microarray analysis of control plant lines that underwent Cre-mediated excision of the ABF3 transgene revealed only two genes that were differentially expressed in more than one plant line, suggesting that the impact of Cre recombinase on the transcriptome was minimal. In the absence of drought stress, overexpression of ABF3 had no effect on the transcriptome, but following drought stress, differences were observed in the gene expression patterns of plants overexpressing ABF3 relative to control plants. Examination of the functional distribution of the differentially expressed genes revealed strong similarity indicating that unintended pathways were not activated. In response to drought stress, overexpression of ABF3 results in a reprogramming of the drought response, which is characterized by changes in the timing or strength of expression of some drought response genes, without activating any unexpected gene networks. These results illustrate that important gene networks are highly regulated in Arabidopsis and that engineering stress tolerance may not necessarily cause extensive changes to the transcriptome.
Project description:In this series of experiments, we wanted to study the transcriptional responses of plants to different levels of water limitation. For mild drought stress, we controlled water potential (a scientific concept for dryness of soil) by using an automated watering system. This system adds water to soil based on the pF values reported by a pF sensor in soil. pF is a classical and widely used index of water potential that was first defined by Schofield (1935). For severe stress conditions, we withheld watering or even dried plants on a lab bench. Here, the transcriptional profiles were compared between well-watered and Md2-treated rice seedling shoots.<br>Md2 (Mild drought level 2): We designated Md2 as a term that means the level of drought severity corresponding to pF2.5, which is equal to a soil matric potential of -31.0 kPa. pF2.5: The value of pF shows the soil matric potential of -31.0 kPa. pF is an index that is equivalent to the effort required for plant root systems to extract water stored in soil.