Mapping the castor bean endosperm proteome revealed a metabolic interaction between plastid, mitochondria, and peroxisomes to optimize seedling growth.

In this work, we studied castor-oil plant Ricinus communis as a classical system for endosperm reserve breakdown. The seeds of castor beans consist of a centrally located embryo with the two thin cotyledons surrounded by the endosperm. The endosperm functions as major storage tissue and is packed with nutritional reserves, such as oil, proteins, and starch. Upon germination, mobilization of the storage reserves requires inter-organellar interplay of plastids, mitochondria, and peroxisomes to optimize growth for the developing seedling. To understand their metabolic interactions, we performed a large-scale organellar proteomic study on castor bean endosperm. Organelles from endosperm of etiolated seedlings were isolated and subjected to liquid chromatography-tandem mass spectrometry (LC-MS/MS). Computer-assisted deconvolution algorithms were applied to reliably assign the identified proteins to their correct subcellular localization and to determine the abundance of the different organelles in the heterogeneous protein samples. The data obtained were used to build a comprehensive metabolic model for plastids, mitochondria, and peroxisomes during storage reserve mobilization in castor bean endosperm.

iTRAQ based proteomic analysis of rice lines having single or stacked blast resistance genes: Pi54/Pi54rh during incompatible interaction with Magnaporthe oryzae.

Deployment of single or multiple blast resistance (R) genes in rice plant is considered to be the most promising approach to enhance resistance against blast disease caused by fungus Magnaporthe oryzae. At the proteome level, relatively little information about R gene mediated defence mechanisms for single and stacking resistance characteristics is available. The overall objective of this study is to look at the proteomics of rice plants that have R genes; Pi54, Pi54rh and stacked Pi54 + Pi54rh in response to rice blast infection. In this study 'isobaric tag for relative and absolute quantification' (iTRAQ)-based proteomics analysis was performed in rice plants at 72-h post inoculation with Magnaporthe oryzae and various differentially expressed proteins were identified in these three transgenic lines in comparison to wild type during resistance response to blast pathogen. Through STRING analysis, the observed proteins were further examined to anticipate their linked partners, and it was shown that several defense-related proteins were co-expressed. These proteins can be employed as targets in future rice resistance breeding against Magnaporthe oryzae. The current study is the first to report a proteomics investigation of rice lines that express single blast R gene Pi54, Pi54rh and stacked (Pi54 + Pi54rh) during incompatible interaction with Magnaporthe oryzae. The differentially expressed proteins indicated that secondary metabolites, reactive oxygen species-related proteins, phenylpropanoid, phytohormones and pathogenesis-related proteins have a substantial relationship with the defense response against Magnaporthe oryzae. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-023-01327-3.

Potential of Trichoderma harzianum and Its Metabolites to Protect Wheat Seedlings against Fusarium culmorum and 2,4-D.

Wheat is a critically important crop. The application of fungi, such as Trichoderma harzianum, to protect and improve crop yields could become an alternative solution to synthetic chemicals. However, the interaction between the fungus and wheat in the presence of stress factors at the molecular level has not been fully elucidated. In the present work, we exposed germinating seeds of wheat (Triticum aestivum) to the plant pathogen Fusarium culmorum and the popular herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) in the presence of T. harzianum or its extracellular metabolites. Then, the harvested roots and shoots were analyzed using spectrometry, 2D-PAGE, and MALDI-TOF/MS techniques. Although F. culmorum and 2,4-D were found to disturb seed germination and the chlorophyll content, T. harzianum partly alleviated these negative effects and reduced the synthesis of zearalenone by F. culmorum. Moreover, T. harzianum decreased the activity of oxidoreduction enzymes (CAT and SOD) and the contents of the oxylipins 9-Hode, 13-Hode, and 13-Hotre induced by stress factors. Under the influence of various growth conditions, changes were observed in over 40 proteins from the wheat roots. Higher volumes of proteins and enzymes performing oxidoreductive functions, such as catalase, ascorbate peroxidase, cytochrome C peroxidase, and Cu/Zn superoxide dismutase, were found in the Fusarium-inoculated and 2,4-D-treated wheat roots. Additionally, observation of the level of 12-oxo-phytodienoic acid reductase involved in the oxylipin signaling pathway in wheat showed an increase. Trichoderma and its metabolites present in the system leveled out the mentioned proteins to the control volumes. Among the 30 proteins examined in the shoots, the expression of the proteins involved in photosynthesis and oxidative stress response was found to be induced in the presence of the herbicide and the pathogen. In summary, these proteomic and metabolomic studies confirmed that the presence of T. harzianum results in the alleviation of oxidative stress in wheat induced by 2,4-D or F. culmorum.

Trichoderma harzianum metabolites disturb Fusarium culmorum metabolism: Metabolomic and proteomic studies.

Trichoderma species are well known for producing various secondary metabolites in response to different fungal pathogens. This paper reports the effects of the metabolites produced during one-day cultivation of Trichoderma harzianum on the growth and development of the popular pathogen Fusarium culmorum. Inhibition of the growth of the pathogen and production of secondary metabolites including zearalenone was observed on Petri dishes. The presence of proteins such as cytochrome c oxidase subunit 4, glutathione-independent glyoxalase HSP31, and putative peroxiredoxin pmp20 in the extract-treated culture indicated oxidative stress, which was confirmed by the presence of a higher amount of catalase and dismutase in the later hours of the culture. A larger amount of enolase and glyceraldehyde 3-phosphate dehydrogenase resulted in faster growth, and the overexpression of stress protein and Woronin body major protein indicated the activation of defense mechanisms. In addition, a cardinal reduction in major mycotoxin production was noted.

FBPAII and rpoBC, the Two Novel Secreted Proteins Identified by the Proteomic Approach from a Comparative Study between Antibiotic-Sensitive and Antibiotic-Resistant Helicobacter pylori-Associated Gastritis Strains.

Helicobacter pylori infection is the leading cause of chronic gastritis, which can develop into gastric cancer. Eliminating H. pylori infection with antibiotics achieves the prevention of gastric cancer. Currently, the prevalence of H. pylori resistance to clarithromycin and metronidazole, and the dual resistance to metronidazole and clarithromycin (C_R, M_R, and C/M_R, respectively), remains at a high level worldwide. As a means of exploring new candidate proteins for the management of H. pylori infection, secreted proteins from antibiotic-susceptible and antibiotic-resistant H. pylori-associated gastritis strains were obtained by in-solution tryptic digestion coupled with nano-liquid chromatography tandem mass spectrometry (nano-LC-MS/MS). A total of 583, 582, 590, and 578 differential expressed proteins were identified from C_R, M_R, C/M_R, and antibiotic-sensitive strain (S_S) samples, respectively. Of these, 23 overlapping proteins were found by Venn diagram analysis. Based on heat map analyses, the most and least differing protein expressions were observed from C/M_R strains and S_S strains, respectively. Of the proteins secreted by the S_S strain, only nine were found. After predicting the protein interaction with metronidazole and clarithromycin via the STITCH database, the two most interesting proteins were found to be rpoBC and FBPAII. After quantitative real-time reverse transcription PCR (qRT-PCR) analysis, a downregulation of rpoB from M_R strains was observed, suggesting a relationship of rpoB to metronidazole sensitivity. Inversely, an upregulation of fba from C_R, M_R, and C/M_R strains was noticed, suggesting the paradoxical expression of FBPAII and the fba gene. This report is the first to demonstrate the association of these two novel secreted proteins, namely, rpoBC and FBPAII, with antibiotic-sensitive H. pylori-associated gastritis strains.

Beneficial Effects of Baduanjin Exercise on Left Ventricular Remodelling in Patients after Acute Myocardial Infarction: an Exploratory Clinical Trial and Proteomic Analysis.

BACKGROUND: The beneficial effects of physical exercise on cardiac remodelling improvement after myocardial infarction have already been suggested. However, the results of previous clinical trials have not been consistent. Moreover, the putative molecular mechanisms leading to the clinically observed effects of physical exercise still remain elusive. AIM: We aimed to evaluate whether the well-defined and strictly controlled traditional Chinese Qigong Baduanjin exercise (BE) would attenuate the adverse left ventricular (LV) remodelling in patients with ST-elevation myocardial infarction (STEMI). METHODS: A total of 110 clinically stable STEMI patients, following successful revascularization of their infarcted coronary arteries, were randomized and enrolled in two groups: 56 were subjected to a 12-week BE-based cardiac rehabilitation programme (BE group), and the remaining 54 were exposed to the usual physical exercise (control group) for the same time period. The primary outcome was the change from baseline to 6 months in the echocardiographic LV end-diastolic volume index (ΔLVEDVi). Proteomic analysis was also performed to uncover associated mechanisms. RESULTS: Compared with the control group, the BE group showed significantly lower ΔLVEDVi (-5.1 ± 1.1 vs. 0.3 ± 1.2 mL/m2, P < 0.01). Proteomic analysis revealed BE-induced variations in the expression of 80 proteins linked to regulation the of metabolic process, immune process, and extracellular matrix reorganization. Furthermore, correlation analyses between the validated serum proteomes and primary endpoint demonstrated a positive association between ΔLVEDVi and MMP-9 expression, but a negative correlation between ΔLVEDVi and CXCL1 expression. CONCLUSION: This is the first study indicating that BE in STEMI patients can alleviate adverse LV remodelling associated with beneficial energy metabolism adaptation, inflammation curbing, and extracellular matrix organization adjustment.

Location of neonatal microglia drives small extracellular vesicles content and biological functions in vitro.

Combining proteomics and systems biology approaches, we demonstrate that neonatal microglial cells derived from two different CNS locations, cortex and spinal cord, and cultured in vitro displayed different phenotypes upon different physiological or pathological conditions. These cells also exhibited greater variability in terms of cellular and small extracellular vesicles (sEVs) protein content and levels. Bioinformatic data analysis showed that cortical microglia exerted anti-inflammatory and neurogenesis/tumorigenesis properties, while the spinal cord microglia were more inflammatory. Interestingly, while both sEVs microglia sources enhanced growth of DRGs processes, only the spinal cord-derived sEVs microglia under LPS stimulation significantly attenuated glioma proliferation. These results were confirmed using the neurite outgrowth assay on DRGs cells and glioma proliferation analysis in 3D spheroid cultures. Results from these in vitro assays suggest that the microglia localized at different CNS regions can ensure different biological functions. Together, this study indicates that neonatal microglia locations regulate their physiological and pathological functional fates and could affect the high prevalence of brain vs spinal cord gliomas in adults.

Proteomics study of rapamycin-injured human coronary artery endothelial cells based on two dimension fluorescence differential gel electrophoresis / 重庆医学

Objective To explore the molecular mechanisms of the effect of eluting stent drug rapamycin for injuring human coronary artery endothelial cells(HCAECs)by using the proteomics method.Methods HCAECs were treated with rapamycin,and the differentially expressed proteins were analyzed by two dimension fluorescence differential gel electrophoresis(2D-DIGE).The changed proteins were identified by MALDI-ToF-ToF.Results At least 85 differential protein spots were found,including 49 up-regulated and 36 down-regulated protein spots.Twenty-six proteins were identified by MALDI-ToF-ToF,including the endoplasmic reticulum protein,mitochondrial protein,molecular chaperones,ubiquitin system related protein,structural protein and oxidative stress related proteins,etc.Conclusion The changes of specific proteins of HCAECs injury induced by rapamycin are investigated by the proteomic method.

Proteomic study provides new clues for complications of hemodialysis caused by dialysis membrane.

The complications of hemodialysis accompanied the hemodialysis and threaten the patients' life. Besides the loss of nutrient substance, such as amino acid and vitamin, we found new clues that the adsorbed proteins on common-used polysulfone-based dialysis membrane might be the reason according to the qualitative proteomic study by ionic liquid assisted sample preparation method. Our results indicated that the adsorbed proteins on the membrane were related with complement activation, blood coagulation, and leukocyte-related biological process. The quantitative proteome further demonstrated some significant changes of signal proteins in the post-dialysis plasma after the hemodialysis, such as beta-2-microglobulin and platelet factor-4, which would further verify these new clues.

Effect of Coenzyme Q10 on Proteomic Profile of Rat Brain Amygdala during Acute Metabolic Stress.

Differences in the proteomic profiles of the brain amygdala in rats with different prognostic resistance to stress were found on the model of metabolic stress. Differential expression of tropomodulin-2, GTP-binding protein SAR1, peroxiredoxin-2, calcineurin B homologous protein 1, Ras-related protein Rab-14, glutathione S-transferase omega-1, Tcrb protein, and NADH dehydrogenase [ubiquinone] iron-sulfur protein 8 (mitochondrial) was shown to depend on the behavioral pattern of animals and stage of the study. Specific features were observed in the involvement of the amygdala in the stress response of specimens with various behavioral characteristics.

Topics in Study Design and Analysis for Multistage Clinical Proteomics Studies.

This chapter discusses the design issues in clinical proteomics study and provides specific suggestions for addressing these questions when using the standard guidelines for the planning. It provides two methods for the sample size estimation in study design. The first method is used for the planning of a clinical proteomic study at the discovery or verification stage; the second method is proposed for the systematic planning of a multistage study. The second part of the chapter introduces three approaches to analyzing the clinical proteomic study and provides analyses for two case studies of clinical proteomic discoveries.

Combined Plasma and Tissue Proteomic Study of Atherogenic Model Mouse: Approach To Elucidate Molecular Determinants in Atherosclerosis Development.

Atherogenic cardiovascular diseases are the major cause of mortality. Prevention and prediction of incidents is important; however, biomarkers that directly reflect the disease progression remain poorly investigated. To elucidate molecular determinants of atherogenesis, proteomic approaches are advantageous by using model animals for comparing changes occurring systematically (bloodstream) and locally (lesion) in accordance with the disease progression stages. We conducted differential mass spectrometric analysis between apolipoprotein E deficient (apoED) and wild-type (wt) mice using the plasma and arterial tissue of both types of mice obtained at four pathognomonic time points of the disease. A total of 100 proteins in the plasma and 390 in the arterial tissues were continuously detected throughout the four time points; 29 were identified in common. Of those, 13 proteins in the plasma and 36 in the arterial tissues showed significant difference in abundance between the apoED and wt mice at certain time points. Importantly, we found that quantitative variation patterns regarding the pathognomonic time points did not always correspond between the plasma and arterial tissues, resulting in gaining insight into atherosclerotic plaque progression. These characteristic proteins were found to be components of inflammation, thrombus formation, and vascular remodeling, suggesting drastic and integrative alteration in accordance with atherosclerosis development.

Enzyme immunoassay and proteomic characterization of troponin I as a marker of mammalian muscle compounds in raw meat and some meat products.

The skeletal muscle protein troponin I (TnI) has been characterized as a potential thermally stable and species-specific biomarker of mammalian muscle tissues in raw meat and meat products. This study proposed a technique for the quantification of TnI comprising protein extraction and sandwich enzyme-linked immunosorbent assay (ELISA). The technique is characterized by a TnI detection limit of 4.8 ng/ml with quantifiable concentrations ranging from 8.7 to 52 ng/ml. The method was shown to be suitable for detection of TnI in mammalian (beef, pork, lamb, and horse) meat but not in poultry (chicken, turkey, and duck) meat. In particular, the TnI content in beef was 0.40 3 ± 0.058 mg/g of wet tissue. The TnI estimations obtained for the pork and beef samples using ELISA were comparable to the proteomic analysis results. Thus, the quantitative study of TnI can be a convenient way to assess the mammalian muscle tissue content of various meat products.

Proteomic analysis of enzyme production by Bacillus licheniformis using different feather wastes as the sole fermentation media.

This study evaluates the use of different types of feathers as fermentation media for enzyme production. Bacillus licheniformis was grown on the feathers, which lead to total biodegradation due to bacterial enzymatic hydrolytic excretion. B. licheniformis excretes protease and lipase activity, with feather concentration being the main parameter controlling their generation. Using a proteomic approach, the proteins excreted during fermentation were identified, and the influence of the chemical composition of the feathers on protein secretion was tested. The identified proteins are hydrolytic enzymes such as keratinase, gamma-glutamyltranspeptidase, chitosanases, and glicosidases. The diversity of proteins is related to the chemical complexity of the feathers. Understanding the composition of a hydrolytic system, when B. licheniformis is cultured on different feathers, may assist in utilizing such a system for producing different hydrolytic enzymes. The data indicate that proteomics can be a valuable tool for describing the physiological state of B. licheniformis cell populations growing on different wastes.

Comparative proteomics analysis of chronic atrophic gastritis: changes of protein expression in chronic atrophic gastritis without Helicobacter pylori infection

Chronic atrophic gastritis (CAG) is a very common gastritis and one of the major precursor lesions of gastric cancer, one of the most common cancers worldwide. The molecular mechanism underlying CAG is unclear, but its elucidation is essential for the prevention and early detection of gastric cancer and appropriate intervention. A combination of two-dimensional gel electrophoresis and mass spectrometry was used in the present study to analyze the differentially expressed proteins. Samples from 21 patients (9 females and 12 males; mean age: 61.8 years) were used. We identified 18 differentially expressed proteins in CAG compared with matched normal mucosa. Eight proteins were up-regulated and 10 down-regulated in CAG when compared with the same amounts of proteins in individually matched normal gastric mucosa. Two novel proteins, proteasome activator subunit 1 (PSME1), which was down-regulated in CAG, and ribosomal protein S12 (RPS12), which was up-regulated in CAG, were further investigated. Their expression was validated by Western blot and RT-PCR in 15 CAG samples matched with normal mucosa. The expression level of RPS12 was significantly higher in CAG than in matched normal gastric mucosa (P < 0.05). In contrast, the expression level of PSME1 in CAG was significantly lower than in matched normal gastric mucosa (P < 0.05). This study clearly demonstrated that there are some changes in protein expression between CAG and normal mucosa. In these changes, down-regulation of PSME1 and up-regulation of RPS12 could be involved in the development of CAG. Thus, the differentially expressed proteins might play important roles in CAG as functional molecules.

Proteomic profiles of mesenchymal stem cells induced by a liver differentiation protocol.

The replacement of disease hepatocytes and the stimulation of endogenous or exogenous regeneration by human mesenchymal stem cells (MSCs) are promising candidates for liver-directed cell therapy. In this study, we isolated MSCs from adult bone marrow by plastic adhesion and induced differentiation with a liver differentiation protocol. Western blot analyses were used to assess the expression of liver-specific markers. Next, MSC-specific proteins were analyzed with two-dimensional (2D) gel electrophoresis and peptide mass fingerprinting matrix-assisted laser desorption/ionization (MALDI)-time of flight (TOF)-mass spectrometry (MS). To confirm the results from the proteomic study, semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) analyses were performed. We demonstrated that MSCs treated with the liver differentiation protocol expressed significantly more albumin, CK19 and CK20, than did undifferentiated cells. In addition the results of proteomic study demonstrated increases expression of FEM1B, PSMC2 and disulfide-isomerase A3 in MSCs treated with the liver differentiation protocol. These results from proteomic profiling will not only provide insight into the global responses of MSCs to hepatocyte differentiation, but will also lead to in-depth studies on the mechanisms of proteomic changes in MSCs.