MTAP-related increased erythroblast proliferation as a mechanism of polycythaemia vera

AbstractPolycythaemia vera (PV) is a haematological disorder caused by an overproduction of erythroid cells. To date, the molecular mechanisms involved in the disease pathogenesis are still ambiguous. This study aims to identify aberrantly expressed proteins in erythroblasts of PV patients by utilizing mass spectrometry-based proteomic analysis. Haematopoietic stem cells (HSCs) were isolated from newly-diagnosed PV patients, PV patients who have received cytoreductive therapy, and healthy subjects. In vitro erythroblast expansion confirmed that the isolated HSCs recapitulated the disease phenotype as the number of erythroblasts from newly-diagnosed PV patients was significantly higher than those from the other groups. Proteomic comparison revealed 17 proteins that were differentially expressed in the erythroblasts from the newly-diagnosed PV patients compared to those from healthy subjects, but which were restored to normal levels in the patients who had received cytoreductive therapy. One of these proteins was S-methyl-5′-thioadenosine phosphorylase (MTAP), which had reduced expression in PV patients’ erythroblasts. Furthermore, MTAP knockdown in normal erythroblasts was shown to enhance their proliferative capacity. Together, this study identifies differentially expressed proteins in erythroblasts of healthy subjects and those of PV patients, indicating that an alteration of protein expression in erythroblasts may be crucial to the pathology of PV.

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PhyllanthusSuppresses Prostate Cancer Cell, PC-3, Proliferation and Induces Apoptosis through Multiple Signalling Pathways (MAPKs, PI3K/Akt, NFκB, and Hypoxia)

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2013/609581 ◽

2013 ◽

Vol 2013 ◽

pp. 1-13 ◽

Cited By ~ 12

Author(s):

Yin-Quan Tang ◽

InduBala Jaganath ◽

Rishya Manikam ◽

Shamala Devi Sekaran

Keyword(s):

Molecular Mechanisms ◽

Prostate Cancer Cell ◽

Prostate Adenocarcinoma ◽

Signalling Pathways ◽

Differentially Expressed Proteins ◽

Signalling Molecules ◽

Anticancer Properties ◽

Anticancer Effects ◽

2D Gel

Phyllanthusis a traditional medicinal plant that has been found to have antihepatitis, antibacterial, and anticancer properties. The present studies were to investigate thein vitromolecular mechanisms of anticancer effects ofPhyllanthus(P. amarus, P. niruri, P. urinaria,andP. watsonii) plant extracts in human prostate adenocarcinoma. The cancer ten-pathway reporter array was performed and revealed that the expression of six pathway reporters were significantly decreased (Wnt, NFκB, Myc/Max, hypoxia, MAPK/ERK, and MAPK/JNK) in PC-3 cells after treatment withPhyllanthusextracts. Western blot was conducted and identified several signalling molecules that were affected in the signalling pathways including pan-Ras, c-Raf, RSK, Elk1, c-Jun, JNK1/2, p38 MAPK, c-myc, DSH,β-catenin, Akt, HIF-1α, GSK3β, NFκB p50 and p52, Bcl-2, Bax, and VEGF, in treated PC-3 cells. A proteomics-based approach, 2D gel electrophoresis, was performed, and mass spectrometry (MS/MS) results revealed that there were 72 differentially expressed proteins identified in treated PC-3 cells and were involved in tumour cell adhesion, apoptosis, glycogenesis and glycolysis, metastasis, angiogenesis, and protein synthesis and energy metabolism. Overall, these findings suggest thatPhyllanthuscan interfere with multiple signalling cascades involved in tumorigenesis and be used as a potential therapeutic candidate for treatment of cancer.

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Identification of Differentially Expressed Proteins in Sugarcane in Response to Infection by Xanthomonas albilineans Using iTRAQ Quantitative Proteomics

Microorganisms ◽

10.3390/microorganisms8010076 ◽

2020 ◽

Vol 8 (1) ◽

pp. 76

Author(s):

Jian-Yu Meng ◽

Mbuya Sylvain Ntambo ◽

Philippe C. Rott ◽

Hua-Ying Fu ◽

Mei-Ting Huang ◽

...

Keyword(s):

Molecular Mechanisms ◽

Vascular System ◽

Lipid Transfer Protein ◽

Differentially Expressed ◽

Post Inoculation ◽

Differentially Expressed Proteins ◽

Lipid Transfer ◽

Leaf Scald ◽

Improvement Programs ◽

Xanthomonas Albilineans

Sugarcane can suffer severe yield losses when affected by leaf scald, a disease caused by Xanthomonas albilineans. This bacterial pathogen colonizes the vascular system of sugarcane, which can result in reduced plant growth and plant death. In order to better understand the molecular mechanisms involved in the resistance of sugarcane to leaf scald, a comparative proteomic study was performed with two sugarcane cultivars inoculated with X. albilineans: one resistant (LCP 85-384) and one susceptible (ROC20) to leaf scald. The iTRAQ (isobaric tags for relative and absolute quantification) approach at 0 and 48 h post-inoculation (hpi) was used to identify and annotate differentially expressed proteins (DEPs). A total of 4295 proteins were associated with 1099 gene ontology (GO) terms by GO analysis. Among those, 285 were DEPs during X. albilineans infection in cultivars LCP 85-384 and ROC20. One hundred seventy-two DEPs were identified in resistant cultivar LCP 85-384, and 113 of these proteins were upregulated and 59 were downregulated. One hundred ninety-two DEPs were found in susceptible cultivar ROC20 and half of these (92) were upregulated, whereas the other half corresponded to downregulated proteins. The significantly upregulated DEPs in LCP 85-384 were involved in metabolic pathways, the biosynthesis of secondary metabolites, and the phenylpropanoid biosynthesis pathway. Additionally, the expression of seven candidate genes related to photosynthesis and glycolytic pathways, plant innate immune system, glycosylation process, plant cytochrome P450, and non-specific lipid transfer protein was verified based on transcription levels in sugarcane during infection by X. albilineans. Our findings shed new light on the differential expression of proteins in sugarcane cultivars in response to infection by X. albilineans. The identification of these genes provides important information for sugarcane variety improvement programs using molecular breeding strategies.

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Comparative iTRAQ proteomics revealed proteins associated with lobed fin regeneration in Bichirs

Proteome Science ◽

10.1186/s12953-019-0153-0 ◽

2019 ◽

Vol 17 (1) ◽

Author(s):

Suxiang Lu ◽

Qian Xiong ◽

Kang Du ◽

Xiaoni Gan ◽

Xuzhen Wang ◽

...

Keyword(s):

Molecular Mechanisms ◽

Multiple Reaction Monitoring ◽

Limb Regeneration ◽

Differentially Expressed ◽

Receptor Interaction ◽

Differentially Expressed Proteins ◽

Absolute Quantitation ◽

Bioinformatics Analyses ◽

Fin Regeneration ◽

Early Stages

Abstract Background Polypterus senegalus can fully regenerate its pectoral lobed fins, including a complex endoskeleton, with remarkable precision. However, despite the enormous potential of this species for use in medical research, its regeneration mechanisms remain largely unknown. Methods To identify the differentially expressed proteins (DEPs) during the early stages of lobed fin regeneration in P. senegalus, we performed a differential proteomic analysis using isobaric tag for relative and absolute quantitation (iTRAQ) approach based quantitative proteome from the pectoral lobed fins at 3 time points. Furthermore, we validated the changes in protein expression with multiple-reaction monitoring (MRM) analysis. Results The experiment yielded a total of 3177 proteins and 15,091 unique peptides including 1006 non-redundant (nr) DEPs. Of these, 592 were upregulated while 349 were downregulated after lobed fin amputation when compared to the original tissue. Bioinformatics analyses showed that the DEPs were mainly associated with Ribosome and RNA transport, metabolic, ECM-receptor interaction, Golgi and endoplasmic reticulum, DNA replication, and Regulation of actin cytoskeleton. Conclusions To our knowledge, this is the first proteomic research to investigate alterations in protein levels and affected pathways in bichirs’ lobe-fin/limb regeneration. In addition, our study demonstrated a highly dynamic regulation during lobed fin regeneration in P. senegalus. These results not only provide a comprehensive dataset on differentially expressed proteins during the early stages of lobe-fin/limb regeneration but also advance our understanding of the molecular mechanisms underlying lobe-fin/limb regeneration.

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Transcriptome profiling of peanut (Arachis hypogaea) gynophores in gravitropic response

Functional Plant Biology ◽

10.1071/fp13075 ◽

2013 ◽

Vol 40 (12) ◽

pp. 1249 ◽

Cited By ~ 7

Author(s):

Hai-fen Li ◽

Xiao-Ping Chen ◽

Fang-he Zhu ◽

Hai-Yan Liu ◽

Yan-Bin Hong ◽

...

Keyword(s):

Differentially Expressed Genes ◽

Arachis Hypogaea ◽

Molecular Mechanisms ◽

Carbon Fixation ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Pentose Phosphate ◽

Differentially Expressed ◽

Pcr Analysis

Peanut (Arachis hypogaea L.) produces flowers aerially, but the fruit develops underground. This process is mediated by the gynophore, which always grows vertically downwards. The genetic basis underlying gravitropic bending of gynophores is not well understood. To identify genes related to gynophore gravitropism, gene expression profiles of gynophores cultured in vitro with tip pointing upward (gravitropic stimulation sample) and downward (control) at both 6 and 12 h were compared through a high-density peanut microarray. After gravitropic stimulation, there were 174 differentially expressed genes, including 91 upregulated and 83 downregulated genes at 6 h, and 491 differentially expressed genes including 129 upregulated and 362 downregulated genes at 12 h. The differentially expressed genes identified were assigned to 24 functional categories. Twenty pathways including carbon fixation, aminoacyl-tRNA biosynthesis, pentose phosphate pathway, starch and sucrose metabolism were identified. The quantitative real-time PCR analysis was performed for validation of microarray results. Our study paves the way to better understand the molecular mechanisms underlying the peanut gynophore gravitropism.

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

Journal of Biomedicine and Biotechnology ◽

10.1155/2010/571343 ◽

2010 ◽

Vol 2010 ◽

pp. 1-11 ◽

Cited By ~ 12

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.

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Differentially Expressed and Novel Transcripts in Highly Purified Chronic Phase CML Stem Cells

Blood ◽

10.1182/blood.v112.11.193.193 ◽

2008 ◽

Vol 112 (11) ◽

pp. 193-193

Author(s):

Yun Zhao ◽

Allen Delaney ◽

Afshin Raouf ◽

Kamini Raghuram ◽

Haiyan I Li ◽

...

Keyword(s):

Stem Cells ◽

Blood Cells ◽

Molecular Mechanisms ◽

Chronic Phase ◽

Differentially Expressed ◽

Pcr Analysis ◽

Direct Role ◽

Hematopoietic Stem ◽

Transcript Levels

Abstract The chronic phase of CML is sustained by rare BCR-ABL+ stem cells. These cells share many properties with normal pluripotent hematopoietic stem cells, but also differ in critical ways that alter their growth, drug responsiveness and genome stability. Understanding the molecular mechanisms underlying the biological differences between normal and CML stem cells is key to the development of more effective CML therapies. To obtain new insights into these mechanisms, we generated Long Serial Analysis of Gene Expression (SAGE) libraries from paired isolates of highly purified lin-CD34+CD45RA-CD36- CD71-CD7-CD38+ and lin-CD34+CD45RA-CD36-CD71-CD7-CD38- cells from 3 chronic phase CML patients (all with predominantly Ph+/BCR-ABL+ cells in both subsets) and from 3 control samples: a pool of 10 normal bone marrows (BMs), a single normal BM and a pool of G-CSF-mobilized blood cells from 9 donors. In vitro bioassays showed the CD34+CD38+ cells were enriched in CFCs (CML: 3–20% pure; normal: 4–19% pure) and the CD34+CD38- cells were enriched in LTC-ICs (CML: 0.2–26% pure; normal: 12–52% pure). Each of the 12 libraries was then sequenced to a depth of ~200,000 tags and tags from libraries prepared from like phenotypes were compared between genotypes using DiscoverySpace software and hierarchical clustering. 1687 (355 with clustering) and 1258 (316 with clustering) transcripts were thus identified as differentially expressed in the CML vs control CD34+CD38− and CD34+CD38+ subsets, respectively. 266 of these transcripts (11 with clustering) were differentially expressed in both subsets. The differential expression of 5 genes (GAS2, IGF2BP2, IL1R1, DUSP1 & SELL) was confirmed by real-time PCR analysis of lin-CD34+ cells isolated from an additional 5 normal BMs and 11 CMLs, and lin-CD34+CD38− cells from an additional 2 normal BMs and 2 CMLs (with dominant Ph+ cells). GAS2 and IL1R1 transcript levels were correlated with BCR-ABL transcript levels in both primitive subsets, and predicted differences in expression of IL1R1 and SELL were apparent within 3 days in CD34+ cord blood cells transduced with a lenti-BCR-ABL-IRES-GFP vs a control lenti-GFP vector (n=3). These findings support a direct role of BCR-ABL in perturbing the expression of these 3 genes. Further comparison of the meta CD34+CD38− and CD34+CD38+ CML cell libraries with most publicly accessible SAGE data revealed 69 novel tags in the CD34+ CML cells that correspond to unique but conserved genomic sequences. Nine of these were recovered by 5′- and 3′- RACE applied to cDNAs pooled from several human leukemic cell lines. These results illustrate the power of SAGE to reveal key components of the transcriptomes of rare human CML stem cell populations including transcripts of genes not previously known to exist. Continuing investigation of their biological roles in primary CML cells and primitive BCR-ABL-transduced human cells offer important strategies for delineating their potential as therapeutic targets.

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

Reproduction ◽

10.1530/rep-13-0313 ◽

2014 ◽

Vol 147 (3) ◽

pp. 321-330 ◽

Cited By ~ 50

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.

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Quantitative Proteomics Analysis to Study the Protective Effects of Nerve Growth Factor on Hippocampal Mitochondria in VD Rats

10.21203/rs.3.rs-456969/v1 ◽

2021 ◽

Author(s):

Yanmei Zhang ◽

yuan yao ◽

Runxiu Zhu ◽

Niyang Aida ◽

Jun Yuan ◽

...

Keyword(s):

Nerve Growth Factor ◽

Growth Factor ◽

Molecular Mechanism ◽

Molecular Mechanisms ◽

Clinical Syndrome ◽

Differentially Expressed ◽

Protective Effects ◽

Differentially Expressed Proteins ◽

Sprague Dawley ◽

Nerve Growth

Abstract Background Vascular dementia (VD) is a kind of clinical syndrome characterized with the impairment cognitive function caused by cerebrovascular disease. Genetics, biochemical, and morphological analyses of cell and animal models, reveal that mitochondria could have roles in this neurodegeneration. Methods We used Sprague-Dawley rats to establish VD model, and used the proteomics method based on relative quantification (iTRAQ) to identify the differentially expressed proteins in hippocampus mitochondria. Results A total of 33 differentially expressed proteins were identified between the VD rats and the VD rats treated with nerve growth factor groups. And five differentially expressed proteins (Rgs14, Slc7a14, Ppm1l, Kcnj10 and Syngr1) were identified after completing the sham-operate control, VD rats and VD rats treated with nerve growth factor groups, then successfully confirmed by western blot. Bioinformatics analysis suggested that the mitochondrial molecular mechanism of VD and the protective effect of nerve growth factor on mitochondrial function of VD rats may be due to different molecular mechanisms. Conclusion We estimated that mitochondrial dysfunction may be the onset of VD and key role in the pathological process of VD. This study not only has a deeper understanding of the mitochondrial molecular mechanism of VD, but also is helpful for the screening of drug targets.

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Comparative iTRAQ Proteomics Identified Myocardium Proteins Associated with Hypoxia of Yak

Current Proteomics ◽

10.2174/1570164616666190123151619 ◽

2019 ◽

Vol 16 (4) ◽

pp. 314-329

Author(s):

Asma Babar ◽

Tserang Donko Mipam ◽

Shixin Wu ◽

Chuanfei Xu ◽

Mujahid Ali Shah ◽

...

Keyword(s):

Protein Expression ◽

High Altitude ◽

Molecular Mechanisms ◽

Pressure Overload ◽

Differentially Expressed ◽

Matrix Remodeling ◽

Cardiovascular Development ◽

Differentially Expressed Proteins ◽

Absolute Quantitation ◽

Hypoxic Adaptation

Background: Yaks inhabit high-altitude are well-adapted to the hypoxic environments. Though, the mechanisms involved in regulatory myocardial protein expression at high-altitude were not completely understood. Objective: To revel the molecular mechanism of hypoxic adaptation in yak, here we have applied comparative myocardial proteomics in between yak and cattle by isobaric Tag for Relative and Absolute Quantitation (iTRAQ) labelling. Methods: To understand the systematic protein expression variations in myocardial tissues that explain the hypoxic adaptation in yak, we have performed iTRAQ analysis combined with Liquid Chromatography- Tandem Mass Spectrometry (LC-MS/MS). Bioinformatics analysis was performed to find the association of these Differentially Expressed Proteins (DEPs) in different functions and pathways. Protein to protein interaction was analyzed by using STRING database. Results: 686 Differentially Expressed Proteins (DEPs) were identified in yak with respect to cattle. From which, 480 DEPs were up-regulated and 206 were down-regulated in yak. Upregulated expression of ASB4, STAT, HRG, RHO and TSP4 in yak may be associated with angiogenesis, cardiovascular development, response to pressure overload to heart and regulation of myocardial contraction in response to increased oxygen tension. The up-regulation of mitochondrial proteins, ACAD8, GPDH-M, PTPMT1, and ALDH2, may have contributed to oxidation within mitochondria, hypoxia-induced cell metabolism and protection of heart against cardiac ischemic injuries. Further, the upregulated expression of SAA1, PTX, HP and MBL2 involved in immune response potentially helpful in myocardial protection against ischemic injuries, extracellular matrix remodeling and free heme neutralization/ clearance in oxygen-deficient environment. Conclusion: Therefore, the identification of these myocardial proteins in will be conducive to investigation of the molecular mechanisms involved in hypoxic adaptations of yaks at high-altitude condition.

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Comparative iTRAQ Proteomic Analysis Provides the Molecular Basis of the Metabolites in Cultivated Glycyrrhiza uralensis and its Wild-Type

Current Proteomics ◽

10.2174/1570164617666191004114511 ◽

2020 ◽

Vol 17 (5) ◽

pp. 365-378

Author(s):

Chengcheng Wang ◽

Lihong Chen ◽

Zhichen Cai ◽

Sijing Feng ◽

Moyi Yue ◽

...

Keyword(s):

Secondary Metabolite ◽

Molecular Mechanisms ◽

Glycyrrhiza Uralensis ◽

Differentially Expressed ◽

Primary Metabolism ◽

Differentially Expressed Proteins ◽

Metabolite Formation ◽

Glycyrrhiza Uralensis Fisch ◽

Significant Difference ◽

Metabolite Synthesis

Background: Licorice is an herbal medicine applied extensively worldwide, and most of the licorice for clinical consumption is provided by Glycyrrhiza uralensis Fisch. Evidence suggests that there is a significant difference in the metabolite composition of licorice from different ecotypes. Objective: To better understand the proteomic changes and molecular mechanisms of metabolite formation in wild and cultivated Glycyrrhiza uralensis Fisch. Methods: Firstly, we established a proteome database by annotating protein sequences according to the genomic and transcriptomic data of G. uralensis. Then, iTRAQ and LC-MS/MS were applied to detect significant protein changes between cultivated and wild G. uralensis. A total of 2751 validated proteins were obtained with high confidence, and 333 were differentially expressed. Differentially expressed proteins were identified and analysed by GO, KEGG, and STRING for network and pathway enrichment. Ultimately, we combined the iTRAQ results with our previous investigation on metabolites to understand the molecular mechanisms underlying metabolite accumulation. Results: The results showed that differentially expressed proteins were mainly involved in the anabolism of carbohydrates and important amino acids that participate in primary metabolism and secondary metabolite synthesis. Another important pathway is the synthesis of flavonoids, which are generally accepted as important bioactive constituents of G. uralensis, and the accumulation of flavonoids in different synthesis stages in two ecotypes of G. uralensis was diverse. Therefore, the differentially abundant proteins in wild and cultivated G. uralensis possibly resulted in differences in medicinal compounds. Conclusion: Our study will provide novel clues for revealing the molecular mechanism of secondary metabolite synthesis as well as quality formation in wild and cultivated G. uralensis.

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