Lipid changes after leaf wounding in Arabidopsis thaliana: expanded lipidomic data form the basis for lipid co‐occurrence analysis

A direct‐infusion electrospray ionization triple–quadrupole mass spectrometry method with multiple reaction monitoring (MRM) was employed to measure 264 lipid analytes extracted from leaves of Arabidopsis thaliana subjected to mechanical wounding. The method provided precise measurements with an average coefficient of variation of 6.1%. Lipid classes analyzed comprised galactolipids and phospholipids (including monoacyl molecular species, molecular species with oxidized acyl chains, phosphatidic acids (PAs)), tri‐ and tetra‐galactosyldiacylglycerols (TrGDGs and TeGDGs), head‐group‐acylated galactolipids, and head‐group‐acylated phosphatidylglycerol (acPG), sulfoquinovosyldiacylglycerols (SQDGs), sphingolipids, di‐ and tri‐acylglycerols (DAGs and TAGs), and sterol derivatives. Of the 264 lipid analytes, 254 changed significantly in response to wounding. In general, levels of structural lipids decreased, whereas monoacyl molecular species, galactolipids and phosphatidylglycerols (PGs) with oxidized fatty acyl chains, PAs, TrGDGs, TeGDGs, TAGs, head‐group‐acylated galactolipids, acPG, and some sterol derivatives increased, many transiently. The observed changes are consistent with activation of lipid oxidizing, hydrolyzing, glycosylating, and acylating activities in the wounding response. Correlation analysis of the levels of lipid analytes across individual control and treated plants was used to construct a lipid dendrogram and to define clusters and sub‐clusters of lipid analytes, each composed of a group of lipids which occurred in a coordinated manner. Current knowledge of metabolism supports the notion that observed sub‐clusters comprise lipids generated by a common enzyme and/or metabolically downstream of a common enzyme. This work demonstrates that co‐occurrence analysis, based on correlation of lipid levels among plants, is a powerful approach to defining lipids generated in vivo by a common enzymatic pathway.     

Phosphatidic acid is a major phospholipid class in reproductive organs of Arabidopsis thaliana

Phospholipids are the crucial components of biological membranes and signal transduction. Among different tissues, flower phospholipids are one of the least characterized features of plant lipidome. Here, we report that floral reproductive organs of Arabidopsis thaliana contain high levels of phosphatidic acid (PA), a known lipid second messenger. By using floral homeotic mutants enriched with specific floral organs, lipidomics study showed increased levels of PA species in ap3-3 mutant with enriched pistils. Accompanied gene expression study for 7 diacylglycerol kinases and 11 PA phosphatases revealed distinct floral organ specificity, suggesting an active phosphorylation/dephosphorylation between PA and diacylglycerol in flowers. Our results suggest that PA is a major phospholipid class in floral reproductive organs of A. thaliana.     

机械损伤对拟南芥莲座叶芥子油苷含量和组成的影响

植物可以利用体内次生代谢产物的变化来抵御昆虫取食和机械损伤.芥子油苷是拟南芥的主要次生代谢产物.通过剪刀剪取叶片(40%面积)对温室培养的拟南芥幼苗莲座叶进行机械损伤处理,观察机械损伤后8个时间点拟南芥叶片中不同种类芥子油苷含量和组合模式的变化.结果表明机械损伤后3 h叶片中芥子油苷总含量开始明显上升,脂肪族和吲哚族芥子油苷含量在损伤后3 h也都显著高于损伤前.在检测到的12种芥子油苷中,4-甲基亚磺酰丁基芥子油苷(4-methylsulphinylbutyl GS,4MSOB)的含量最多,占芥子油苷总量的48.5%,并且在损伤3 h后含量增加.4MSOB含量的变化成为影响莲座叶中芥子油苷组合模式的主导因素.其它各种芥子油苷在损伤后不同时间点的变化也存在差异.     

Homology modeling of Ferredoxin-nitrite reductase from Arabidopsis thaliana

Different subtypes of Influenza A virus are associated with species specific, zoonotic or pandemic Influenza. The cause of its severity underlies in complicated evolution of its segmented RNA genome. Although genetic shift and genetic drift are well known in the evolution of this virus, we reported the significant role of unique RNA palindromes in its evolution. Our computational approach identified the existence of unique palindromes in each subtype of Influenza A virus with its absence in Influenza B relating the fact of virulence and vigorous genetic hitchhiking in Influenza A. The current study focused on the re-assortment event responsible for the emergence of pandemic-2009 H1N1 virus, which is associated with outgrow of new palindrome and in turn, changing its RNA structure. We hypothesize that the change in RNA structure due to the presence of palindrome facilitates the event of re-assortment in Influenza A. Thus the evolutionary process of Influenza A is much more complicated as previously known, and that has been demonstrated in this study.     

The recombination landscape in Arabidopsis thaliana F2 populations

Recombination during meiosis shapes the complement of alleles segregating in the progeny of hybrids, and has important consequences for phenotypic variation. We examined allele frequencies, as well as crossover (XO) locations and frequencies in over 7000 plants from 17 F(2) populations derived from crosses between 18 Arabidopsis thaliana accessions. We observed segregation distortion between parental alleles in over half of our populations. The potential causes of distortion include variation in seed dormancy and lethal epistatic interactions. Such a high occurrence of distortion was only detected here because of the large sample size of each population and the number of populations characterized. Most plants carry only one or two XOs per chromosome pair, and therefore inherit very large, non-recombined genomic fragments from each parent. Recombination frequencies vary between populations but consistently increase adjacent to the centromeres. Importantly, recombination rates do not correlate with whole-genome sequence differences between parental accessions, suggesting that sequence diversity within A. thaliana does not normally reach levels that are high enough to exert a major influence on the formation of XOs. A global knowledge of the patterns of recombination in F(2) populations is crucial to better understand the segregation of phenotypic traits in hybrids, in the laboratory or in the wild.     

Mutations in retrotransposon AtCOPIA4 compromises resistance to Hyaloperonospora parasitica in Arabidopsis thaliana

Retrotransposons (RTEs) are a principal component of most eukaryotic genomes, representing 50%-80% of some grass genomes. RTE sequences have been shown to be preferentially present in disease resistance gene clusters in plants. Arabidopsis thaliana has over 1,600 annotated RTE sequences and 56 of these appear to be expressed because of the exact expressed sequence tag (EST) matches and the presence of intact open reading frames. Of the 22 represented in the Affymetrix ATH1 array, AtCOPIA4 was found to be expressed at a higher level than all other RTEs across different developmental stages. Since AtCOPIA4 is located in the RPP5 gene cluster and is adjacent to RPP4 which confers resistance to the downy mildew oomycete Hyaloperonospora parasitica isolate EMWA1, we evaluated AtCOPIA4 mutants for resistance to this pathogen. T-DNA insertional and antisense knockout of AtCOPIA4 was found to reduce the resistance of wild type plants by 2-4 folds. Our results suggest that retrotransposon can be exapted to participate in plant defense response.     

Spatial and temporal analysis of the local response to wounding

We studied the local response to wounding in Arabidopsis thaliana leaves using a two-step microarray analysis. A microarray containing 3500 cDNA clones was first screened to enrich for genes affected by wounding in the immediate vicinity of the wound (4 h post wounding). 359 non-redundant putative wound responsive genes were then spotted on a smaller wound-response array for detailed analysis of spatial expression (local, adjacent and systemic), timing of expression (0.5, 4, 8, 17 h), and effect of hormone treatments (methyl jasmonate, ethylene and abscisic acid). Our results show that genes that respond early at the site of the wound also respond throughout the plant, with similar kinetics. Early-induced genes which respond systemically encode predominantly signal transduction and regulatory factors (36%), and the expression of many of them is also controlled by methyl jasmonate (about 35% of the 36%). Genes specific to the wound site and the wounded leaf have a slower response to wounding and are mainly metabolic genes. At the wound, many genes of the lignin biosynthesis pathway were induced. In silico analysis of the 5′ promoter regions of genes affected by wounding revealed G-box-related motifs in a significant proportion of the promoters. These results show that the establishment of a systemic response to wounding is a priority for the plant, and that the local response at the wound site is established later. Ethylene and abscisic acid are involved in the local response, regulating repression of photosynthetic genes and expression of drought responsive genes respectively.     

Microarray-based analysis of stress-regulated microRNAs in Arabidopsis thaliana

High-salinity, drought, and low temperature are three common environmental stress factors that seriously influence plant growth and development worldwide. Recently, microRNAs (miRNAs) have emerged as a class of gene expression regulators that have also been linked to stress responses. However, the relationship between miRNA expression and stress responses is just beginning to be explored. Here, we identified 14 stress-inducible miRNAs using microarray data in which the effects of three abiotic stresses were surveyed in Arabidopsis thaliana. Among them, 10 high-salinity-, four drought-, and 10 cold-regulated miRNAs were detected, respectively. miR168, miR171, and miR396 responded to all of the stresses. Expression profiling by RT-PCR analysis showed great cross-talk among the high-salinity, drought, and cold stress signaling pathways. The existence of stress-related elements in miRNA promoter regions provided further evidence supporting our results. These findings extend the current view about miRNA as ubiquitous regulators under stress conditions.     

QTL detection power of multi-parental RIL populations in Arabidopsis thaliana

A major goal of today's biology is to understand the genetic basis of quantitative traits. This can be achieved by statistical methods that evaluate the association between molecular marker variation and phenotypic variation in different types of mapping populations. The objective of this work was to evaluate the statistical power of quantitative trait loci (QTL) detection of various multi-parental mating designs, as well as to assess the reasons for the observed differences. Our study was based on an empirical data of 20 Arabidopsis thaliana accessions, which have been selected to capture the maximum genetic diversity. The examined mating designs differed strongly with respect to the statistical power to detect QTL. We observed the highest power to detect QTL for the diallel cross with random mating design. The results of our study suggested that performing sibling mating within subpopulations of joint-linkage mapping populations has the potential to considerably increase the power for QTL detection. Our results, however, revealed that using designs in which more than two parental alleles segregate in each subpopulation increases the power even more.     

Universally occurring phenylpropanoid and species-specific indolic metabolites in infected and uninfected Arabidopsis thaliana roots and leaves

A total of eleven alkali-released, aromatic compounds were identified by HPLC, MS and NMR analyses in cell wall extracts from Arabidopsis thaliana roots. Nine of them together constituted the three complete series of 4-hydroxy-, 4-hydroxy-3-methoxy, and 4-hydroxy-3,5-dimethoxy-substituted benzaldehydes, benzoic acids and cinnamic acids. The other two were indolic metabolites: indole-3-carboxylic acid and indole-3-carbaldehyde. Qualitatively similar, but quantitatively distinct profiles were obtained using cell-wall extracts from A. thaliana leaves. Several of these compounds, particularly indole-3-carboxylic acid, 4-hydroxybenzoic acid and all four aldehydes, increased considerably in concentration upon infection of roots with Pythium sylvaticum, as did at least some of them upon infection of leaves with Pseudomonas syringae pv tomato. Comparison of these results with analogous data on a variety of different plant species suggests a remarkable structural uniformity among the majority of constitutive as well as infection-induced, aromatic cell wall-bound compounds throughout the entire plant kingdom-in sharp contrast to the highly species-specific, chemically highly divers bouquets of soluble aromatic metabolites.     

Mild ammonium stress increases chlorophyll content in Arabidopsis thaliana

Nitrate (NO₃⁻) and ammonium (NH₄⁺) are the main forms of nitrogen available in the soil for plants. Excessive NH₄⁺ accumulation in tissues is toxic for plants and exclusive NH₄⁺-based nutrition enhances this effect. Ammonium toxicity syndrome commonly includes growth impairment, ion imbalance and chlorosis among others. In this work, we observed high intraspecific variability in chlorophyll content in 47 Arabidopsis thaliana natural accessions grown under 1 mM NH₄⁺ or 1 mM NO₃⁻ as N-source. Interestingly, chlorophyll content increased in every accession upon ammonium nutrition. Moreover, this increase was independent of ammonium tolerance capacity. Thus, chlorosis seems to be an exclusive effect of severe ammonium toxicity while mild ammonium stress induces chlorophyll accumulation.     

ZEITLUPE positively regulates hypocotyl elongation at warm temperature under light in Arabidopsis thaliana

Hypocotyl cell elongation has been studied as a model to understand how cellular expansion contributes to plant organ growth. Hypocotyl elongation is affected by multiple environmental factors, including light quantity and light quality. Red light inhibits hypocotyl growth via the phytochrome signaling pathways. Proteins of the FLAVIN-BINDING KELCH REPEAT F-BOX 1 / LOV KELCH PROTEIN 2 / ZEITLUPE family are positive regulators of hypocotyl elongation under red light in Arabidopsis. These proteins were suggested to reduce phytochrome-mediated inhibition of hypocotyl elongation. Here, we show that ZEITLUPE also functions as a positive regulator in warmth-induced hypocotyl elongation under light in Arabidopsis.     

Intracellular ca2+ stores could participate to abscisic acid-induced depolarization and stomatal closure in Arabidopsis thaliana

In Arabidopsis thaliana cell suspension,abscisic acid (aBa) induces changes in cytosolic calcium concentration ([Ca²⁺]_cyt) which are the trigger for aBa-induced plasma membrane anion current activation, H⁺-aTPase inhibition, and subsequent plasma membrane depolarization. In the present study, we took advantage of this model to analyze the implication of intracellular Ca²⁺ stores in aBa signal transduction through electrophysiological current measurements, cytosolic Ca²⁺ activity measurements with the apoaequorin Ca²⁺ reporter protein and external pH measurement. Intracellular Ca²⁺ stores involvement was determined by using specific inhibitors of CICR channels: the cADP-ribose/ryanodine receptor (Br-cADPR and dantrolene) and of the inositol trisphosphate receptor ({"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122). In addition experiments were performed on epidermal strips of A. thaliana leaves to monitor stomatal closure in response to ABA in presence of the same pharmacology. Our data provide evidence that ryanodine receptor and inositol trisphosphate receptor could be involved in ABA-induced (1) Ca²⁺ release in the cytosol, (2) anion channel activation and H⁺-ATPase inhibition leading to plasma membrane depolarization and (3) stomatal closure. Intracellular Ca²⁺ release could thus contribute to the control of early events in the ABA signal transduction pathway in A. thaliana.     

A systems biology approach to analyse leaf carbohydrate metabolism in Arabidopsis thaliana

Plant carbohydrate metabolism comprises numerous metabolite interconversions, some of which form cycles of metabolite degradation and re-synthesis and are thus referred to as futile cycles. In this study, we present a systems biology approach to analyse any possible regulatory principle that operates such futile cycles based on experimental data for sucrose (Scr) cycling in photosynthetically active leaves of the model plant Arabidopsis thaliana. Kinetic parameters of enzymatic steps in Scr cycling were identified by fitting model simulations to experimental data. A statistical analysis of the kinetic parameters and calculated flux rates allowed for estimation of the variability and supported the predictability of the model. A principal component analysis of the parameter results revealed the identifiability of the model parameters. We investigated the stability properties of Scr cycling and found that feedback inhibition of enzymes catalysing metabolite interconversions at different steps of the cycle have differential influence on stability. Applying this observation to futile cycling of Scr in leaf cells points to the enzyme hexokinase as an important regulator, while the step of Scr degradation by invertases appears subordinate.