Proteomic analysis of Arabidopsis thaliana (L.) Heynh responses to a generalist sucking pest (Myzus persicae Sulzer)

Herbivorous insects can cause severe cellular changes to plant foliage following infestations, depending on feeding behaviour. Here, a proteomic study was conducted to investigate the influence of green peach aphid (Myzus persicae Sulzer) as a polyphagous pest on the defence response of Arabidopsis thaliana (L.) Heynh after aphid colony establishment on the host plant (3 days). Analysis of about 574 protein spots on 2‐DE gels revealed 31 differentially expressed protein spots. Twenty out of these 31 differential proteins were selected for analysis by mass spectrometry. In 12 of the 20 analysed spots, we identified seven and nine proteins using MALDI‐TOF‐MS and LC‐ESI‐MS/MS, respectively. Of the analysed spots, 25% contain two proteins. Different metabolic pathways were modulated in Arabidopsis leaves according to aphid feeding: most corresponded to carbohydrate, amino acid and energy metabolism, photosynthesis, defence response and translation. This paper has established a survey of early alterations induced in the proteome of Arabidopsis by M. persicae aphids. It provides valuable insights into the complex responses of plants to biological stress, particularly for herbivorous insects with sucking feeding behaviour.     

Targeting novel chemical and constitutive primed metabolites against Plectosphaerella cucumerina

Priming is a physiological state for protection of plants against a broad range of pathogens, and is achieved through stimulation of the plant immune system. Various stimuli, such as beneficial microbes and chemical induction, activate defense priming. In the present study, we demonstrate that impairment of the high‐affinity nitrate transporter 2.1 (encoded by NRT2.1) enables Arabidopsis to respond more quickly and strongly to Plectosphaerella cucumerina attack, leading to enhanced resistance. The Arabidopsis thaliana mutant lin1 (affected in NRT2.1) is a priming mutant that displays constitutive resistance to this necrotroph, with no associated developmental or growth costs. Chemically induced priming by β–aminobutyric acid treatment, the constitutive priming mutant ocp3 and the constitutive priming present in the lin1 mutant result in a common metabolic profile within the same plant–pathogen interactions. The defense priming significantly affects sugar metabolism, cell‐wall remodeling and shikimic acid derivatives levels, and results in specific changes in the amino acid profile and three specific branches of Trp metabolism, particularly accumulation of indole acetic acid, indole‐3–carboxaldehyde and camalexin, but not the indolic glucosinolates. Metabolomic analysis facilitated identification of three metabolites in the priming fingerprint: galacturonic acid, indole‐3–carboxylic acid and hypoxanthine. Treatment of plants with the latter two metabolites by soil drenching induced resistance against P. cucumerina, demonstrating that these compounds are key components of defense priming against this necrotrophic fungus. Here we demonstrate that indole‐3–carboxylic acid induces resistance by promoting papillae deposition and H₂O₂ production, and that this is independent of PR1, VSP2 and PDF1.2 priming.     

A Dual‐Purpose Bromocoumarin Tag Enables Deep Profiling of the Cellular Cysteinome

Chemical proteomics enables comprehensive profiling of small molecules in complex proteomes. A critical component to understand the interactome of a small molecule is the precise location on a protein where the interaction takes place. Several approaches have been developed that take advantage of bio‐orthogonal chemistry and subsequent enrichment steps to isolate peptides modified by small molecules. These methods rely on target identification at the level of mass spectrometry making it difficult to interpret an experiment when modified peptides are not identified. Herein, an approach in which fluorescence‐triggered two‐dimensional chromatography enables the isolation of small molecule‐conjugated peptides prior to mass spectrometry analysis is described. In this study, a bromocoumarin moiety has been utilized that fluoresces and generates a distinct isotopic signature to locate and identify modified peptides. Profiling of a cellular cysteinome with the use of a bromocoumarin tag demonstrates that two‐dimensional fluorescence‐based chromatography separation can enable the identification of proteins containing reactive cysteine residues. Moreover, the method facilitates the interrogation of low abundance proteins with greater depth and sensitivity than a previously reported isotope‐targeted approach. Lastly, this workflow enables the identification of small‐molecule modified peptides from a protein‐of‐interest.     

Differential protein expression profile in gastrointestinal stromal tumors

Summary. Gastrointestinal stromal tumors (GISTs) arise from the interstitial cells of Cajal through gain of function mutations of the oncogene KIT. Imatinib offers the first effective treatment for patients with GISTs, but the therapeutic outcome strongly depends on the type of KIT mutation. We used ProteinChip technology to investigate whether GISTs with different KIT mutations express different proteins. In total, 154 proteins were significantly differentially expressed in GISTs with exon 9 KIT mutation compared to GISTs with exon 11 KIT mutation.     

Wnt signaling loss accelerates the appearance of neuropathological hallmarks of Alzheimer's disease in J20‐APP transgenic and wild‐type mice

Alzheimer's disease (AD ) is a neurodegenerative pathology characterized by aggregates of amyloid‐β (Aβ) and phosphorylated tau protein, synaptic dysfunction, and spatial memory impairment. The Wnt signaling pathway has several key functions in the adult brain and has been associated with AD , mainly as a neuroprotective factor against Aβ toxicity and tau phosphorylation. However, dysfunction of Wnt/β‐catenin signaling might also play a role in the onset and development of the disease. J20 APP swInd transgenic (Tg) mouse model of AD was treated i.p. with various Wnt signaling inhibitors for 10 weeks during pre‐symptomatic stages. Then, cognitive, biochemical and histochemical analyses were performed. Wnt signaling inhibitors induced severe changes in the hippocampus, including alterations in Wnt pathway components and loss of Wnt signaling function, severe cognitive deficits, increased tau phosphorylation and Aβ_1–42 peptide levels, decreased Aβ42/Aβ40 ratio and Aβ_1–42 concentration in the cerebral spinal fluid, and high levels of soluble Aβ species and synaptotoxic oligomers in the hippocampus, together with changes in the amount and size of senile plaques. More important, we also observed severe alterations in treated wild‐type (WT ) mice, including behavioral impairment, tau phosphorylation, increased Aβ_1–42 in the hippocampus, decreased Aβ_1–42 in the cerebral spinal fluid, and hippocampal dysfunction. Wnt inhibition accelerated the development of the pathology in a Tg AD mouse model and contributed to the development of Alzheimer's‐like changes in WT mice. These results indicate that Wnt signaling plays important roles in the structure and function of the adult hippocampus and suggest that inhibition of the Wnt signaling pathway is an important factor in the pathogenesis of AD .

Read the Editorial Highlight for this article on page 356 .

     

Evaluation of tricin,a stylet probing stimulant of brown planthopper,in infested and non‐infested rice plants

The flavone, tricin (5,7,4′‐trihydroxy‐3′,5′‐dimethoxyflavone), is a valuable secondary metabolite that is common in gramineous plants, including cultivated rice (Oryza sativa). It can defend the rice plant against infestation by the brown planthopper (BPH), Nilaparvata lugens Stål, one of the most important pests of rice. This study evaluated the tricin concentration in infested and non‐infested rice plants. The results of the liquid chromatography coupled to tandem mass spectrometry (LC–MS/MS) quantitative analysis showed that the tricin concentration in rice leaves was significantly higher than in the stems and roots. The mass concentration of tricin in the leaves at the leaf stage was significantly higher than at the tiller and booting stages. The relationship between rice variety, BPH resistance and tricin concentration was investigated. There was a significant negative correlation between tricin concentration and the injury severity scores for rice varieties. Moreover, BPH infestation caused variations in tricin concentration among rice plants. High BPH infestation levels can significantly reduce the tricin concentration in rice plants. However, there is no significant effect of the length of infestation times on tricin concentrations in rice leaves. These results suggest that there may be an elicitor in BPH saliva, which is injected into rice plants during BPH infestation and triggers the tricin metabolic system. Future studies need to identify the elicitor and clarify the mechanism underlying tricin reduction in infested rice plants.     

Quick Identification of Piperidine Alkaloid from Roots of Grewia nervosa and Their Glucosidase Inhibitory Activity

Grewia nervosa is a herbal plant used in traditional medicine for different purposes. Bioassay‐guided chemical fractionation of G. nervosa roots resulted in an identification of two known and one new compound, namely microgrewiapine A, homomicrogrewiapine, and N‐methylmicrocosamine, respectively. Their structures were determined using combination of LC/HR‐MS, ¹H‐NMR, and IR spectral analyses and followed by comparison with those reported in the literature. The problematic separation of these alkaloids on traditional column chromatography (Silica gel, Octadecyl silane, Sephadex) was resolved by using HPLC. Structurally similar compounds from the piperidine family have been characterized by using HR‐MS analysis in combination with NMR data of crude samples. The major constituent i.e. N‐methylmicrocosamine isolated from the butanol fraction of methanol root extract (MRE) was found to possess the dose dependent α‐glucosidase inhibition activity with an IC₅₀ value of 53.40 μm . Furthermore, N‐methylmicrocosamine showed maximum α‐glucosidase inhibition of 97.48 ± 0.7% at 107.5 μm , which is approximately 1.3 × 10³ fold higher than the activity shown by acarbose (97.72% inhibition at 61.95 mm ), a standard anti‐diabetic drug available commercially. This work also reports the in vitro α‐glucosidase inhibitory activity of the major alkaloids isolated from G. nervosa for the first time.     

NMR,HS‐SPME‐GC/MS,and HPLC/MSn Analyses of Phytoconstituents and Aroma Profile of Rosmarinus eriocalyx

In this work, a comprehensive study on the chemical constituents of the aerial parts of Rosmarinus eriocalyx (Lamiaceae), an aromatic shrub traditionally consumed as a food and herbal remedy in Algeria, is presented. The aroma profile was analysed by headspace solid phase microextraction (HS‐SPME) coupled with gas chromatography‐mass spectrometry (GC/MS), whereas the crude extract constituents were analyzed by ¹H‐NMR and by high performance liquid chromatography coupled with mass spectrometry (HPLC/MSⁿ). Thirty‐nine volatile compounds, most of them being monoterpenes, have been identified, with camphor, camphene, and α‐pinene as the most abundant constituents. ¹H‐NMR analysis revealed the presence of phenolic compounds and betulinic acid while HPLC/MSⁿ allowed the identification of glycosilated and aglyconic flavonoids as well as phenylpropanoid derivatives. Some of these constituents, namely as betulinic acid, rosmanol, and cirsimaritin were reported for the first time in R. eriocalyx.     

UHPLC/HR‐ESI‐MS/MS Profiling of Phenolics from Tunisian Lycium arabicum Boiss. Antioxidant and Anti‐lipase Activities’ Evaluation

This study was performed in the aim to evaluate nine different extracts from Tunisian Lycium arabicum for their total phenolic and total flavonoid contents, phytochemical analyses as well as their antioxidant and anti‐lipase activities. The in vitro antioxidant property was investigated using three complementary methods (DPPH, ferric reducing antioxidant power (FRAP), and β‐carotene‐linoleic acid bleaching assays) while anti‐lipase activity was evaluated using 4‐methylumbelliferyl oleate method. From all of the tested extracts the most potent found to be the polar MeOH extracts especially those of stems and leaves. In order to investigate the chemical composition of these extracts and possible correlation of their constituents with the observed activities, an UHPLC/HR‐ESI‐MS/MS analysis was performed. Several compounds belonging to different chemical classes were tentatively identified such as rutin and kampferol rutinoside, the major constituents of the leaves, and N‐caffeoyltyramine, lyciumide A, N‐dihydrocaffeoyltyramine as well as fatty acids: trihydroxyoctadecadienoic acid and hydroxyoctadecadienoic acid isomers were detected abundantly in the stems. These results showed that the MeOH extracts of stems and leaves of L. arabicum can be considered as a potential source of biological active compounds.     

Chemical Diversity and Antimicrobial Activity of Volatile Compounds from Zanthoxylum zanthoxyloides Lam. according to Compound Classes,Plant Organs and Senegalese Sample Locations

The chemical diversity of Zanthoxylum zanthoxyloides growing wild in Senegal was studied according to volatile compound classes, plant organs and sample locations. The composition of fruit essential oil was investigated using an original targeted approach based on the combination of gas chromatography (GC) and liquid chromatography (LC) both coupled with mass spectrometry (MS). The volatile composition of Z. zanthoxyloides fruits exhibited relative high amounts of hydrocarbon monoterpenes (24.3 – 55.8%) and non‐terpenic oxygenated compounds (34.5 – 63.1%). The main components were (E)‐β‐ocimene (12.1 – 39%), octyl acetate (11.6 – 21.8%) and decanol (9.7 – 15.4%). The GC and GC/MS profiling of fruit essential oils showed a chemical variability according to geographical locations of plant material. The LC/MS/MS analysis of fruit oils allowed the detection of seven coumarins in trace content. The chemical composition of fruit essential oils was compared with volatile fractions of leaves and barks (root and trunk) from the same plant station. Hexadecanoic acid, germacrene D and decanal were identified as the major constituents of leaves whereas the barks (root and trunk) were dominated by pellitorine (85.8% and 57%, respectively), an atypic linear compound with amide group. The fruit essential oil exhibited interesting antimicrobial activities against Staphylococcus aureus and Candida albicans, particularly the alcohol fraction of the oil.