iTRAQ-Based Quantitative Proteomic Analysis of Pseudostellaria heterophylla from Geo-Authentic Habitat and Cultivated Bases

2019 ◽  
Vol 16 (3) ◽  
pp. 231-245
Author(s):  
Yujiao Hua ◽  
Chengcheng Wang ◽  
Shengnan Wang ◽  
Zixiu Liu ◽  
Xunhong Liu ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Large Subunit ◽  
Absolute Quantification ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Realtime Pcr ◽  
Pseudostellaria Heterophylla ◽  
Response To Stress ◽  
Isobaric Tags ◽  
Shock Proteins

Background: Pseudostellaria heterophylla is an important tonic traditional Chinese medicine. However, the molecular changes in the herb from geo-authentic habitat and cultivated bases remain to be explored. Objective: The purpose of this research was to study differences in P. heterophylla from geo-authentic habitat and cultivated bases. Methods: High-throughput technologies of transcriptomic and proteomic were used to identify proteins. Isobaric Tags for Relative and Absolute Quantification (iTRAQ) MS/MS has been utilized to evaluate changes in P. heterophylla from geo-authentic habitat and cultivated bases. Results: In this study, a total of 3775 proteins were detected, and 140 differentially expressed proteins were found in P. heterophylla from geo-authentic habitat and cultivated bases. 44 significantly differential expressed proteins were identified based on functional analysis classified into nine categories. Five differentially expressed proteins were confirmed at the gene expression level by Quantitative realtime PCR. Catabolic metabolism, carbohydrate metabolism, and response to stress of oxidoreductases and transferases in P. heterophylla from geo-authentic habitat were stronger than in those from cultivated bases, but protein folding and response to stress of heat shock proteins, isomerases, rubisco large subunit-binding proteins, chaperone proteins, and luminal-binding proteins in herbs from cultivated bases were more active. ADG1 and TKTA could be the critical proteins to regulate sucrose; MFP2 and CYS may be the crucial proteins that control the metabolism of fatty acids and amino acids. Conclusion: These results will provide the basic information for exploring the differences in secondary metabolites in P. heterophylla from geo-authentic habitat and cultivated bases and the protein mechanism of its quality formation.

scholarly journals Identification of differentially expressed proteins in fresh and frozen–thawed boar spermatozoa by iTRAQ-coupled 2D LC–MS/MS

Reproduction ◽  
2014 ◽  
Vol 147 (3) ◽  
pp. 321-330 ◽  
Author(s):  
Xiaoli Chen ◽  
Huabin Zhu ◽  
Chuanhuo Hu ◽  
Haisheng Hao ◽  
Junfang Zhang ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Bioinformatic Analysis ◽  
Absolute Quantification ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Protein Levels ◽  
Boar Semen ◽  
Isobaric Tags ◽  
Boar Spermatozoa

Cryodamage is a major problem in semen cryopreservation, causing changes in the levels of proteins that influence the function and motility of spermatozoa. In this study, protein samples prepared from fresh and frozen–thawed boar spermatozoa were compared using the isobaric tags for relative and absolute quantification (iTRAQ) labeling technique coupled to 2D LC–MS/MS analysis. A total of 41 differentially expressed proteins were identified and quantified, including 35 proteins that were present at higher levels and six proteins that were present at lower levels in frozen–thawed spermatozoa by at least a mean of 1.79-fold (P<0.05). On classifying into ten distinct categories using bioinformatic analysis, most of the 41 differentially expressed proteins were found to be closely relevant to sperm premature capacitation, adhesions, energy supply, and sperm–oocyte binding and fusion. The expression of four of these proteins, SOD1, TPI1, ODF2, and AKAP3, was verified by western blot analysis. We propose that alterations in these identified proteins affect the quality of cryopreserved semen and ultimately lower its fertilizing capacity. This is the first study to compare protein levels in fresh and frozen–thawed spermatozoa using the iTRAQ technology. Our preliminary results provide an overview of the molecular mechanisms of cryodamage in frozen–thawed spermatozoa and theoretical guidance to improve the cryopreservation of boar semen.

scholarly journals Comparative Proteomics of Schistosoma Japonicum Developed From Different Oncomelania Snails Permissive Areas

2021 ◽  
Author(s):  
Feng Miao ◽  
Yongbin Wang ◽  
Kun Yang ◽  
Wei Li ◽  
Chunrong Xiong ◽  
...  
Keyword(s):  
Absolute Quantification ◽  
Shandong Province ◽  
Receptor Expression ◽  
Differentially Expressed ◽  
Water Diversion ◽  
Differentially Expressed Proteins ◽  
Histone H4 ◽  
Intermediate Hosts ◽  
Male Adult ◽  
Isobaric Tags

Abstract Background: Schistosomiasis is an important zoonotic parasitic disease that is widely prevalent in tropical and subtropical countries and regions in the worldwide.Methods: We aimed to analyze the proteomic differences between adult S. japonicum worms in Weishan Lake of Shandong province and the Jiangsu Yangtse river. Isobaric tags for relative and absolute quantification (iTRAQ) assays were used to analyze the differential proteomic profiles between female and male adult worms.Results: A total of 2364 adult S. japonicum proteins were identified, and 1901 proteins were quantified by isobaric tags for relative and absolute quantification (iTRAQ) technology. Our results revealed 68 differentially expressed proteins (DEPs) in female adult worms and 55 DEPs in male adult worms. LC-MS/MS and bioinformatics analysis indicated that these DEPs are enriched in cellular composition, molecular function, biological function and catabolism pathways. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Domain and Clusters of Orthologous Groups (COG) analyses indicated that several groups of DEPs were involved in regulating transport, metabolism, signal transduction, energy production and conversion, defense and biosynthesis in adult S. japonicum worms. Our findings indicated that adult S. japonicum worms derived from O. hupensis that were transferred from permissive to nonpermissive areas exhibited moderate changes at the proteomic level. Moreover, snails transferred to the Weishan Lake did not change their schistosomiasis transmission ability and remained pathogenic in mice. In addition, three upregulated proteins (peptidylprolyl isomerase, heat shock protein 90α and receptor expression-enhancing protein (Q5DBJ1)) and three downregulated proteins (histone H3, histone H4 and receptor expression-enhancing protein (C1L9D7)) were found in both female and male adult worms. Conclusions: Proteomic analysis showed that differentially expressed proteins (DEPs) between adult S. japonicum worms in Weishan Lake of Shandong province and the Jiangsu Yangtse river. The results of this proteomics analysis of adult worms that hatched in two separate intermediate hosts help to improve our understanding of the growth and developmental mechanisms of S. japonicum in different environments. Under the South-to-North Water Diversion Project (SNWDP) framework, long-term surveillance is needed to prevent the diffusion of O. hupensis and to reduce the risk of schistosomiasis transmission.

scholarly journals Screening for Proteins Related to The Biosynthesis of Hispidin and Its Derivatives in Phellinus Igniarius Using iTRAQ Proteomic Analysis

2020 ◽  
Author(s):  
Jinjing Guo ◽  
Xiaoxi Liu ◽  
Yuanjie Li ◽  
Hongyan Ji ◽  
Cheng Liu ◽  
...  
Keyword(s):  
Stress Responses ◽  
Multiple Reaction Monitoring ◽  
Enrichment Analysis ◽  
Absolute Quantification ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Phellinus Igniarius ◽  
Chemical Structures ◽  
Isobaric Tags

Abstract Background: Hispidin (HIP) and its derivatives, a class of natural fungal metabolites, possess extremely complex and interesting chemical structures with extensive pharmacological activities. Phellinus igniarius, which is the most common sources of HIP, can be used as both medicine and food. The biosynthetic pasthway of HIP in P. igniarius is not yet clear and hence effective regulatory mechanism of HIP is absent.The purpose of this paper was to illustrate a biosynthesis system for hispidin and its derivatives at the protein level. Results: We found that tricetolatone (TL) is a key biosynthetic precursor in the biosynthetic pathway of hispidin and that its addition led to increased production of hispidin and various hispidin derivatives. Based on the changes in the concentrations of precursors and intermediates, key timepoints in the biosynthetic process were identified. We used isobaric tags for relative and absolute quantification (iTRAQ) to study dynamic changes of related proteins in vitro. The 270 differentially expressed proteins were determined by GO enrichment analysis to be primarily related to energy metabolism, oxidative phosphorylation, and environmental stress responses after TL supplementation. The differentially expressed proteins were related to ATP synthase, NAD binding protein, oxidoreductase, and other elements associated with electron transfer and dehydrogenation reactions during the biosynthesis of hispidin and its derivatives. Multiple reaction monitoring (MRM) technology was used to selectively verify the iTRAQ results, leading us to screen 11 proteins that were predicted to be related to the biosynthesis pathways.Conclution: These findings help to clarify the molecular mechanism of biosynthesis of hispidin and its derivatives and may serve as a foundation for future strategies to identify new hispidin derivatives.

scholarly journals iTRAQ Quantitative Analysis of Multidrug Resistance Mechanisms in Human Gastric Cancer Cells

2010 ◽  
Vol 2010 ◽  
pp. 1-11 ◽  
Author(s):  
Huai-Dong Hu ◽  
Feng Ye ◽  
Da-Zhi Zhang ◽  
Peng Hu ◽  
Hong Ren ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Multidrug Resistance ◽  
Cell Line ◽  
Molecular Mechanisms ◽  
Absolute Quantification ◽  
Differentially Expressed ◽  
Human Gastric Cancer ◽  
Differentially Expressed Proteins ◽  
Gastric Cancer Cells ◽  
Proteomic Approach

Multidrug resistance (MDR) is a major obstacle towards a successful treatment of gastric cancer. However, the mechanisms of MDR are intricate and have not been fully understood. To elucidate the molecular mechanisms of MDR in gastric cancer, we employed the proteomic approach of isobaric tags for relative and absolute quantification (iTRAQ), followed by LC-MS/MS, using the vincristine-resistant SGC7901/VCR cell line and its parental SGC7901 cell line as a model. In total, 820 unique proteins were identified and 91 proteins showed to be differentially expressed in SGC7901/VCR compared with SGC7901. Several differentially expressed proteins were further validated by western blot analysis. Furthermore, the association of MVP, one of the highly expressed proteins in SGC7901/VCR, with MDR was verified. Our study is the first application of iTRAQ technology for MDR mechanisms analysis in gastric cancer, and many of the differentially expressed proteins identified have not been linked to MDR in gastric cancer before, which showed the value of this technology in identifying differentially expressed proteins in cancer.

scholarly journals Isobaric Tags for Relative and Absolute Quantitation-Based Quantitative Serum Proteomics Analysis in Ischemic Stroke Patients With Hemorrhagic Transformation

2021 ◽  
Vol 15 ◽  
Author(s):  
Zhifeng Qi ◽  
Shuhua Yuan ◽  
Xixi Zhou ◽  
Xunming Ji ◽  
Ke Jian Liu
Keyword(s):  
Ischemic Stroke ◽  
Interaction Analysis ◽  
Hemorrhagic Transformation ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Stroke Patients ◽  
Absolute Quantitation ◽  
Metabolic Pathway Analysis ◽  
Serum Proteomics ◽  
Isobaric Tags

Hemorrhagic transformation (HT), which occurs with or without reperfusion treatments (thrombolysis and/or thrombectomy), deteriorates the outcomes of ischemic stroke patients. It is essential to find clinically reliable biomarkers that can predict HT. In this study, we screened for potential serum biomarkers from an existing blood bank and database with 243 suspected acute ischemic stroke (AIS) patients. A total of 37 patients were enrolled, who were diagnosed as AIS without receiving reperfusion treatment. They were divided into two groups based on whether they were accompanied with HT or not (five HT and 32 non-HT). Serum samples were labeled by isobaric tags for relative and absolute quantitation (iTRAQ) and analyzed by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and compared under NCBInr database. A total of 647 proteins in sera samples were captured, and the levels of 17 proteins (12 upregulated and five downregulated) were significantly different. These differentially expressed proteins were further categorized with Gene Ontology functional classification annotation and Kyoto Encyclopedia of Genes and Genomes metabolic pathway analysis into biological processes. Further protein–protein interaction analysis using String database discovered that, among the differentially expressed proteins, 10 pairs of proteins were found to have crosstalk connections, which may have direct (physical) and indirect (functional) interactions for the development of HT. Our findings suggest that these differentially expressed proteins could serve as potential biomarkers for predicting HT after ischemic stroke.

scholarly journals Study on Differential Protein Expression in Natural Selenium-Enriched and Non-Selenium-Enriched Rice Based on iTRAQ Quantitative Proteomics

Biomolecules ◽  
2019 ◽  
Vol 9 (4) ◽  
pp. 130 ◽  
Author(s):  
Rui Zeng ◽  
Muhammad Farooq ◽  
Li Wang ◽  
Yang Su ◽  
Tengda Zheng ◽  
...  
Keyword(s):  
Amino Acid ◽  
Differential Expression ◽  
Starch Synthesis ◽  
Oxygen Metabolism ◽  
Absolute Quantification ◽  
Stress Effect ◽  
Cysteine Synthase ◽  
Liquid Chromatography Tandem Mass ◽  
Isobaric Tags ◽  
Shock Proteins

This work was designated to scrutinize the protein differential expression in natural selenium-enriched and non-selenium-enriched rice using the Isobaric-tags for relative and absolute quantification (iTRAQ) proteomics approach. The extracted proteins were subjected to enzyme digestion, desalting, and identified by iTRAQ coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) technology. High pH C18 separation analysis was performed, and the data were then analyzed by Protein PilotTM (V4.5) search engine. Protein differential expression was searched out by comparing relatively quantified proteins. The analysis was conducted using gene ontology (GO), cluster of orthologous groups of proteins (COG) and Kyoto encyclopedia of genes and genomes (KEGG) metabolic pathways. A total of 3235 proteins were detected and 3161 proteins were quantified, of which 401 were differential proteins. 208 down-regulated and 193 up-regulated proteins were unveiled. 77 targeted significant differentially expressed proteins were screened out for further analysis, and were classified into 10 categories: oxidoreductases, transferases, isomerases, heat shock proteins, lyases, hydrolases, ligases, synthetases, tubulin, and actin. The results indicated that the anti-stress, anti-oxidation, active oxygen metabolism, carbohydrate and amino acid metabolism of natural selenium-enriched rice was higher than that of non-selenium rice. The activation of the starch synthesis pathway was found to be bounteous in non-selenium-enriched rice. Cysteine synthase (CYS) and methyltransferase (metE) might be the two key proteins that cause amino acid differences. OsAPx02, CatC, riPHGPX, HSP70 and HSP90 might be the key enzymes regulating antioxidant and anti-stress effect differences in two types of rice. This study provides basic information about deviations in protein mechanism and secondary metabolites in selenium-enriched and non-selenium-enriched rice.

scholarly journals Understanding the Toxin Effects of β-Zearalenol and HT-2 on Bovine Granulosa Cells Using iTRAQ-Based Proteomics

Animals ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 130
Author(s):  
Lian Li ◽  
Min Yang ◽  
Chengmin Li ◽  
Fangxiao Yang ◽  
Genlin Wang
Keyword(s):  
Granulosa Cells ◽  
Absolute Quantification ◽  
P Value ◽  
Toxic Response ◽  
Human Food Chain ◽  
Response Profile ◽  
Isobaric Tags ◽  
Shock Proteins ◽  
Cellular Components ◽  
Marker Molecule

Zearalenone (ZEA) and T-2 are the most common mycotoxins in grains and can enter the animal and human food-chain and cause many health disorders. To elucidate the toxic response profile, we stimulated bovine granulosa cells (GCs) with β-zearalenol or HT-2. Using isobaric tags for relative and absolute quantification (iTRAQ)-based proteomic, 178 and 291 differentially expressed proteins (DEPs, fold change ≥ 1.3 and p-value < 0.05) in β-zearalenol and HT-2 groups were identified, respectively. Among these DEPs, there were 66 common DEPs between β-zearalenol and HT-2 groups. These 66 DEPs were associated with 23 biological processes terms, 14 molecular functions terms, and 19 cellular components terms. Most heat shock proteins (HSPs) were involved in the toxic response. Reactive oxygen species accumulation, the endoplasmic reticulum (ER)-stress related marker molecule (GRP78), and apoptosis were activated. β-zearalenol and HT-2 inhibited oestradiol (E2) production. These results emphasized the important function of HSPs, clarified oxidative stress, and demonstrated the caspase-3 signaling cascade involved in mycotoxin-treated toxic response, along with decreased E2 production. This study offers new insights into the toxicity of β-zearalenol and HT-2 on ovarian granulosa cells.

scholarly journals Integrated Analysis of Transcriptomic and Proteomics Data Reveals the Induction Effects of Rotenoid Biosynthesis of Mirabilis himalaica Caused by UV-B Radiation

2018 ◽  
Vol 19 (11) ◽  
pp. 3324 ◽  
Author(s):  
Li Gu ◽  
Weilie Zheng ◽  
Mingjie Li ◽  
Hong Quan ◽  
Jianming Wang ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Plant Hormone ◽  
Absolute Quantification ◽  
Differentially Expressed ◽  
Integrated Analysis ◽  
Signaling System ◽  
Proteomics Data ◽  
Adaptation Mechanism ◽  
Molecular Events ◽  
Isobaric Tags

Mirabilis himalaica (Edgew.) Heimerl is one of the most important genuine medicinal plants in Tibet, in which the special plateau habitat has been associated with its excellent medicinal quality and efficacy. However, the mechanisms by which environmental factors affect biosynthesis of secondary metabolic components remain unclear in this species. In this study, RNA sequencing and iTRAQ (isobaric Tags for Relative and Absolute Quantification) techniques were used to investigate the critical molecular “events” of rotenoid biosynthesis responding to UV-B radiation, a typical plateau ecological factor presented in native environment-grown M. himalaica plants. A total of 3641 differentially expressed genes (DEGs) and 106 differentially expressed proteins (DEPs) were identified in M. himalaica between UV-B treatment and control check (CK). Comprehensive analysis of protein and transcript data sets resulted in 14 and 7 DEGs from the plant hormone signal transduction and phosphatidylinositol signaling system pathways, respectively, being significantly enriched. The result showed that the plant hormone signal transduction and phosphatidylinositol signaling system might be the key metabolic strategy of UV-B radiation to improve the biosynthesis of rotenoid in M. himalaica. At same time, most of the DEGs were associated with auxin and calcium signaling, inferring that they might drive the downstream transmission of these signal transduction pathways. Regarding those pathways, two chalcone synthase enzymes, which play key roles in the biosynthesis of rotenoid that were thought as the representative medicinal component of M. himalaica, were significantly upregulated in UV-B radiation. This study provides a theoretical basis for further exploration of the adaptation mechanism of M. himalaica to UV-B radiation, and references for cultivation standardization.

scholarly journals iTRAQ-Based Quantitative Proteomic Analysis Reveals Changes in Metabolite Biosynthesis in Monascus purpureus in Response to a Low-Frequency Magnetic Field

Toxins ◽  
2018 ◽  
Vol 10 (11) ◽  
pp. 440 ◽  
Author(s):  
Jialan Zhang ◽  
Yingbao Liu ◽  
Li Li ◽  
Mengxiang Gao
Keyword(s):  
Protein Level ◽  
Low Frequency ◽  
Absolute Quantification ◽  
Differentially Expressed ◽  
Monascus Purpureus ◽  
Monacolin K ◽  
Yellow Orange ◽  
Frequency Magnetic Field ◽  
Isobaric Tags ◽  
Control Protein

Background: Low-frequency magnetic fields (LF-MFs) dampen the citrinin output by Monascus purpureus in fermentations. The influence of LF-MFs on biosynthesis by M. purpureus was evaluated at the protein level. Methods: Cultures were treated with a 1.6-mT MF from day 0 to day 2 of incubation, and secondary metabolite production was evaluated on the day 12 of incubation. All proteins were extracted from M. purpureus mycelia and subjected to isobaric tags for relative and absolute quantification (iTRAQ) labeling and subsequent liquid chromatography/mass spectrometry (LC-MS/MS) analysis on day 6 of fermentation. Results: There was no difference in biomass between the treated samples and the control. Citrinin production was 46.7% lower, and the yields of monacolin K and yellow, orange, and red pigment were 29.3%, 31.3%, 41.7%, and 40.3% higher, respectively, in the exposed samples compared to the control. Protein expression in M. purpureus under LF-MF treatment was quantified using iTRAQ technology. Of 2031 detected proteins, 205 were differentially expressed. The differentially-expressed proteins were subjected to Gene Ontology (GO) functional annotation and statistical analysis, which revealed that they mainly refer to biological metabolism, translation, antioxidant, transport and defense pathways. Among all the tagged proteins, emphasis was placed on the analysis of those involved in the synthesis of citrinin, pigment and monacolin K was emphasized. Conclusions: LF-MFs affected Monascus secondary metabolism at the protein level, and aggregate data for all the protein profiles in LF-MF-treated Monascus was obtained.

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