Proteome Science[JOURNAL]
Sun
200 papers
Proteomic investigation of protein adsorption to cerebral microdialysis membranes in surgically treated intracerebral hemorrhage patients - a pilot study.
Tobieson L, Czifra Z, Wåhlén K … +2 more , Marklund N, Ghafouri B
Proteome Sci · 2020 · PMID 32728348 · Full text
BACKGROUND: Cerebral microdialysis (CMD) is a minimally invasive technique for sampling the interstitial fluid in human brain tissue. CMD allows monitoring the metabolic state of tissue, as well as sampling macromolecule...
BACKGROUND: Cerebral microdialysis (CMD) is a minimally invasive technique for sampling the interstitial fluid in human brain tissue. CMD allows monitoring the metabolic state of tissue, as well as sampling macromolecules such as proteins and peptides. Recovery of proteins or peptides can be hampered by their adsorption to the CMD membrane as has been previously shown in-vitro however, protein adsorption to CMD membranes has not been characterized following implantation in human brain tissue. METHODS: In this paper, we describe the pattern of proteins adsorbed to CMD membranes compared to that of the microdialysate and of cerebrospinal fluid (CSF). We retrieved CMD membranes from three surgically treated intracerebral hemorrhage (ICH) patients, and analyzed protein adsorption to the membranes using two-dimensional gel electrophoresis (2-DE) in combination with nano-liquid mass spectrometry. We compared the proteome profile of three compartments; the CMD membrane, the microdialysate and ventricular CSF collected at time of CMD removal. RESULTS: We found unique protein patterns in the molecular weight range of 10-35 kDa for each of the three compartments. CONCLUSION: This study highlights the importance of analyzing the membranes in addition to the microdialysate when using CMD to sample proteins for biomarker investigation.
Proteome-wide identification of arginine methylation in colorectal cancer tissues from patients.
Lim Y, Lee JY, Ha SJ … +3 more , Yu S, Shin JK, Kim HC
Proteome Sci · 2020 · PMID 32467672 · Full text
BACKGROUND: Protein arginine methylation reaction is catalyzed by protein arginine methyltransferase (PRMT) and the modification is implicated in various diseases including cancer. Currently, thousands of arginine methyl...
BACKGROUND: Protein arginine methylation reaction is catalyzed by protein arginine methyltransferase (PRMT) and the modification is implicated in various diseases including cancer. Currently, thousands of arginine methylation sites have been identified using high-resolution mass spectrometry-based proteomics technology. However, identification of arginine methylation using clinical samples at proteome level has not been reported yet. The objective of the present study was to identify, monomethyl-arginine (MMA) and asymmetric dimethyl-arginine (ADMA) sites in colorectal cancer (CRC) tissues at proteome level. METHODS: Pooled CRC tissue samples from 10 patients with stage II and III were digested by trypsin and these digests were further processed and lyophilized. Using monomethyl- or asymmetric dimethyl arginine (MMA or ADMA, respectively) motif kits, methylarginine-containing peptides were enriched and subsequently analyzed by high-resolution LC-MS/MS. DLD1 and HCT116 colon cancer cells were treated with type I PRMTs inhibitor (MS023) alone or combined with SN-38, and the effect of the drugs on CRC cell proliferation and apoptosis was measured by water-soluble tetrazolium salt (WST-1) assay and FACS analysis, respectively. RESULTS: In the present study, 455 MMA sites of 272 proteins and 314 ADMA sites of 155 proteins were identified from CRC tissues acquired from patients. In addition, 216 methylation sites and 75 substrates for PRMTs were newly identified. These results reveal the significant presence of MMA and ADMA sites on nucleic acid binding proteins and protein complexes involved in transcription. To investigate the effect of protein arginine methylation in CRC proliferation and apoptosis, MS023 was treated to two CRC cell lines. After 48 h treatment with various concentrations of MS023, CRC cell proliferation was significantly suppressed, with concomitant apoptosis induction. Furthermore, MS023 treatment significantly enhanced the inhibitory effect of SN-38 on CRC cell proliferation. CONCLUSION: This work reports the first comprehensive analysis of arginine methylation with clinical sample and suggests that type I PRMTs are potential therapeutic targets for drug discovery in CRC.
Production of high-quality two-dimensional gel electrophoresis profile for marine medaka samples by using Trizol-based protein extraction approaches.
Kwok CS, Lai KK, Lam SW … +3 more , Chan KK, Xu SJ, Lee FW
Proteome Sci · 2020 · PMID 32390769 · Full text
BACKGROUND: Marine medaka is among the most popular models of fish species for ecotoxicology and environmental research and proteomic studies are useful tools for understanding the molecular responses of medaka upon expo...
BACKGROUND: Marine medaka is among the most popular models of fish species for ecotoxicology and environmental research and proteomic studies are useful tools for understanding the molecular responses of medaka upon exposure to different environmental stressors. The preparation of high-quality protein samples is the key to producing high-quality two-dimensional gel electrophoresis (2-DE) results for proteomic analysis. In recent years, Trizol-based protein extraction has been gaining popularity because of its promising performance in producing high-quality 2-DE as well as the convenience of the method. METHODS: Three Trizol-based approaches (Trizol method, Aliquot Trizol method and Trizol method with a commercial clean-up kit) were used to extract proteins from a marine medaka sample and 2-DE profiles were produced. Quality of the 2-DE profiles and effectiveness of the extraction methods were evaluated. For comparison, two common protein extraction methods (lysis buffer method and trichloroacetic acid (TCA)/acetone precipitation extraction) were also applied in parallel to Trizol-based approaches. RESULTS: Any of the three Trizol-based approaches produced a high-quality 2-DE profile of marine medaka compared with both lysis buffer method and TCA/acetone precipitation extraction. In addition, Trizol method with a commercial clean-up kit produced the best 2-DE profile in terms of background clarity, number of spots and resolution of proteins. CONCLUSIONS: Trizol-based approaches offered better choices than traditional protein extraction methods for 2-DE analysis of marine medaka. The modified version of Trizol method with a commercial clean-up kit was shown to produce the best 2-DE profile.
NEK10 interactome and depletion reveal new roles in mitochondria.
Peres de Oliveira A, Basei FL, Slepicka PF … +9 more , de Castro Ferezin C, Melo-Hanchuk TD, de Souza EE, Lima TI, Dos Santos VT, Mendes D, Silveira LR, Menck CFM, Kobarg J
Proteome Sci · 2020 · PMID 32368190 · Full text
BACKGROUND: Members of the family of NEK protein kinases (NIMA-related kinases) were described to have crucial roles in regulating different aspects of the cell cycle. NEK10 was reported to take part in the maintenance o...
BACKGROUND: Members of the family of NEK protein kinases (NIMA-related kinases) were described to have crucial roles in regulating different aspects of the cell cycle. NEK10 was reported to take part in the maintenance of the G2/M checkpoint after exposure to ultraviolet light. NEK1, NEK5, NEK2 and NEK4 proteins on the other hand have been linked to mitochondrial functions. METHODS: HEK293T cells were transfected with FLAG empty vector or FLAG-NEK10 and treated or not with Zeocin. For proteomic analysis, proteins co-precipitated with the FLAG constructs were digested by trypsin, and then analyzed via LC-MS/MS. Proteomic data retrieved were next submitted to Integrated Interactome System analysis and differentially expressed proteins were attributed to Gene Ontology biological processes and assembled in protein networks by Cytoscape. For functional, cellular and molecular analyses two stable Nek10 silenced HeLa cell clones were established. RESULTS: Here, we discovered the following possible new NEK10 protein interactors, related to mitochondrial functions: SIRT3, ATAD3A, ATAD3B, and OAT. After zeocin treatment, the spectrum of mitochondrial interactors increased by the proteins: FKBP4, TXN, PFDN2, ATAD3B, MRPL12, ATP5J, DUT, YWHAE, CS, SIRT3, HSPA9, PDHB, GLUD1, DDX3X, and APEX1. We confirmed the interaction of NEK10 and GLUD1 by proximity ligation assay and confocal microscopy. Furthermore, we demonstrated that NEK10-depleted cells showed more fragmented mitochondria compared to the control cells. The knock down of NEK10 resulted further in changes in mitochondrial reactive oxygen species (ROS) levels, decreased citrate synthase activity, and culminated in inhibition of mitochondrial respiration, affecting particularly ATP-linked oxygen consumption rate and spare capacity. NEK10 depletion also decreased the ratio of mtDNA amplification, possibly due to DNA damage. However, the total mtDNA content increased, suggesting that NEK10 may be involved in the control of mtDNA content. CONCLUSIONS: Taken together these data place NEK10 as a novel regulatory player in mitochondrial homeostasis and energy metabolism.
Protection of the transplant kidney during cold perfusion with doxycycline: proteomic analysis in a rat model.
Moser MAJ, Sawicka K, Sawicka J … +4 more , Franczak A, Cohen A, Bil-Lula I, Sawicki G
Proteome Sci · 2020 · PMID 32336955 · Full text
BACKGROUND: It has been previously shown that doxycycline (Doxy) protects the kidney from preservation injury by inhibition of matrix metalloproteinase. However, the precise molecular mechanism involved in this protectio...
BACKGROUND: It has been previously shown that doxycycline (Doxy) protects the kidney from preservation injury by inhibition of matrix metalloproteinase. However, the precise molecular mechanism involved in this protection from injury is not known. We used a pharmaco-proteomics approach to identify potential molecular targets associated with kidney preservation injury. METHODS: Rat kidneys were cold perfused with or without doxycycline (Doxy) for 22 h. Kidneys perfusates were analyzed for the presence of injury markers such as lactate dehydrogenase (LDH), and neutrophil-gelatinase associated lipocalin (NGAL). Proteins extracted from kidney tissue were analyzed by 2-dimensional gel electrophoresis. Proteins of interest were identified by mass spectrometry. RESULTS: Triosephosphate isomerase, PGM, dihydropteridine reductase-2, pyridine nucleotide-disulfide oxidoreductase, phosphotriesterase-related protein, and aminoacylase-1A were not affected by cold perfusion. Perfusion with Doxy increased their levels. N(G),N(G)-dimethylarginine dimethylaminohydrolase and phosphoglycerate kinase 1 were decreased after cold perfusion. Perfusion with Doxy led to an increase in their levels. CONCLUSIONS: This study revealed specific metabolic enzymes involved in preservation injury and in the mechanism whereby Doxy protects the kidney against injury during cold perfusion.
Comprehensive analysis of the cardiac proteome in a rat model of myocardial ischemia-reperfusion using a TMT-based quantitative proteomic strategy.
Lim SH, Lee J, Han MJ
Proteome Sci · 2020 · PMID 32165865 · Full text
BACKGROUND: Traditional studies of the cardiac proteome have mainly investigated in an animal model by two-dimensional gel electrophoresis (2-DE). However, the results have not been of satisfactory quality for an underst...
BACKGROUND: Traditional studies of the cardiac proteome have mainly investigated in an animal model by two-dimensional gel electrophoresis (2-DE). However, the results have not been of satisfactory quality for an understanding of the underlying mechanism. Recent quantitative proteomic methods have been improved to overcome these limitations. To comprehensively study the cardiac proteome in a rat model of ischemia-reperfusion (IR), we developed a tandem mass tag (TMT)-based quantitative proteomic strategy. Furthermore, using this strategy, we examined the molecular mechanisms underlying the prevention of myocardial infarction by the intake of L. extract (TALE), a representative dietary fiber grain. METHODS: Cardiac proteomes were analyzed by 2-DE as a gel-based approach, and TMT labeling coupled with two-dimensional liquid chromatography (2D-LC) and tandem mass spectrometry (MS/MS) as a non-gel-based quantitative approach. Additionally, gene ontology annotation was conducted by PANTHER database. Several proteins of interest were verified by a Western blot analysis. RESULTS: Total 641 proteins were identified commonly from two independent MS datasets using 2D-LC MS/MS. Among these, we identified 151 IR-related proteins that were differentially expressed between the sham-operation group and IR group, comprising 62 up-regulated proteins and 89 down-regulated proteins. Most of the reduced proteins were involved in metabolic processes. In addition, 57 of the IR-related proteins were affected by TALE intake, representing 25 up-regulated proteins and 32 down-regulated proteins. In particular, TALE intake leads to a switch in metabolism to reduce the loss of high-energy phosphates and the accumulation of harmful catabolites (especially reactive oxygen species (ROS)) and to maintain cytoskeleton balance, leading to a reduction in cardiac IR injury. CONCLUSIONS: Our study provides a comprehensive proteome map of IR-related proteins and potential target proteins and identifies mechanisms implicated in the prevention of myocardial infarction by TALE intake in a rat IR model.
Comparative proteomic analysis of the brain and colon in three rat models of irritable bowel syndrome.
Zhang B, Xue H, Wang W … +7 more , Chen T, Su M, Kang N, Yang J, Bian Z, Wang F, Tang X
Proteome Sci · 2020 · PMID 32123521 · Full text
BACKGROUND: Irritable bowel syndrome (IBS) has been gradually recognized as a disorder of the brain-gut interaction, but the molecular changes in the brain and colon that occur in disease development remain poorly unders...
BACKGROUND: Irritable bowel syndrome (IBS) has been gradually recognized as a disorder of the brain-gut interaction, but the molecular changes in the brain and colon that occur in disease development remain poorly understood. We employed proteomic analysis to identify differentially expressed proteins in both the brain and colon of three IBS models. METHODS: To explore the relevant protein abundance changes in the brain and colon, isobaric tags for relative and absolute quantitation (iTRAQ), liquid chromatography and tandem mass spectrometry (LC-MS) and Western blotting methods were used in three IBS models, including maternal separation (MS, group B), chronic wrap restraint stress (CWRS, group C) and a combination of MS and CWRS (group D). RESULTS: We identified 153, 280, and 239 proteins that were common and differentially expressed in the two tissue types of groups B, C and D, respectively; 43 differentially expressed proteins showed the same expression changes among the three groups, including 25 proteins upregulated in the colon and downregulated in the brain, 7 proteins downregulated in the colon and upregulated in the brain, and 3 proteins upregulated and 8 downregulated in both tissues. Gene ontology analysis showed that the differentially expressed proteins were mainly associated with cellular assembly and organization and cellular function and maintenance. Protein interaction network and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that the differentiated proteins were mainly involved in the protein ubiquitination pathway and mitochondrial dysfunction. CONCLUSIONS: Taken together, the data presented represent a comprehensive and quantitative proteomic analysis of the brain and colon in IBS models, providing new evidence of an abnormal brain-gut interaction in IBS. These data may be useful for further investigation of potential targets in the diagnosis and treatment of IBS.
The interactome and proteomic responses of ALKBH7 in cell lines by in-depth proteomics analysis.
Meng S, Zhan S, Dou W … +1 more , Ge W
Proteome Sci · 2019 · PMID 31889914 · Full text
BACKGROUND: ALKBH7 is a mitochondrial protein, involved in programmed necrosis, fatty acid metabolism, cell cycle regulation, and prostate cancer disease. However, the exact roles of ALKBH7 and the underlying molecular m...
BACKGROUND: ALKBH7 is a mitochondrial protein, involved in programmed necrosis, fatty acid metabolism, cell cycle regulation, and prostate cancer disease. However, the exact roles of ALKBH7 and the underlying molecular mechanisms remain mysterious. Thus, investigations of the interactome and proteomic responses of ALKBH7 in cell lines using proteomics strategies are urgently required. METHODS: In the present study, we investigated the interactome of ALKBH7 in mitochondria through immunoprecipitation-mass spectrometry/mass spectrometry (IP-MS/MS). Additionally, we established the ALKBH7 knockdown and overexpression cell lines and further identified the differentially expressed proteins (DEPs) in these cell lines by TMT-based MS/MS. Two DEPs (UQCRH and HMGN1) were validated by western blotting analysis. RESULTS: Through bioinformatic analysis the proteomics data, we found that ALKBH7 was involved in protein homeostasis and cellular immunity, as well as cell proliferation, lipid metabolism, and programmed necrosis by regulating the expression of PTMA, PTMS, UQCRH, HMGN1, and HMGN2. Knockdown of ALKBH7 resulted in upregulation of UQCRH and HMGN1 expression, and the opposite pattern of expression was detected in ALKBH7 overexpression cell lines; these results were consistent with our proteomics data. CONCLUSION: Our findings indicate that the expression of UQCRH and HMGN1 is regulated by ALKBH7, which provides potential directions for future studies of ALKBH7. Furthermore, our results also provide comprehensive insights into the molecular mechanisms and pathways associated with ALKBH7.
Chronic kidney disease: a review of proteomic and metabolomic approaches to membranous glomerulonephritis, focal segmental glomerulosclerosis, and IgA nephropathy biomarkers.
Taherkhani A, Farrokhi Yekta R, Mohseni M … +2 more , Saidijam M, Arefi Oskouie A
Proteome Sci · 2019 · PMID 31889913 · Full text
Chronic Kidney Disease (CKD) is a global health problem annually affecting millions of people around the world. It is a comprehensive syndrome, and various factors may contribute to its occurrence. In this study, it was...
Chronic Kidney Disease (CKD) is a global health problem annually affecting millions of people around the world. It is a comprehensive syndrome, and various factors may contribute to its occurrence. In this study, it was attempted to provide an accurate definition of chronic kidney disease; followed by focusing and discussing on molecular pathogenesis, novel diagnosis approaches based on biomarkers, recent effective antigens and new therapeutic procedures related to high-risk chronic kidney disease such as membranous glomerulonephritis, focal segmental glomerulosclerosis, and IgA nephropathy, which may lead to end-stage renal diseases. Additionally, a considerable number of metabolites and proteins that have previously been discovered and recommended as potential biomarkers of various CKD using '-omics-' technologies, proteomics, and metabolomics were reviewed.
Comparative iTRAQ proteomics revealed proteins associated with lobed fin regeneration in Bichirs.
Lu S, Xiong Q, Du K … +6 more , Gan X, Wang X, Yang L, Wang Y, Ge F, He S
Proteome Sci · 2019 · PMID 31832023 · Full text
BACKGROUND: 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 regeneratio...
BACKGROUND: 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 , 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 . 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.
Exploring the key communicator role of exosomes in cancer microenvironment through proteomics.
Kim H, Kim DW, Cho JY
Proteome Sci · 2019 · PMID 31686989 · Full text
There have been many attempts to fully understand the mechanism of cancer behavior. Yet, how cancers develop and metastasize still remain elusive. Emerging concepts of cancer biology in recent years have focused on the c...
There have been many attempts to fully understand the mechanism of cancer behavior. Yet, how cancers develop and metastasize still remain elusive. Emerging concepts of cancer biology in recent years have focused on the communication of cancer with its microenvironment, since cancer cannot grow and live alone. Cancer needs to communicate with other cells for survival, and thus they secrete various messengers, including exosomes that contain many proteins, miRNAs, mRNAs, etc., for construction of the tumor microenvironment. Moreover, these intercellular communications between cancer and its microenvironment, including stromal cells or distant cells, can promote tumor growth, metastasis, and escape from immune surveillance. In this review, we summarized the role of proteins in the exosome as communicators between cancer and its microenvironment. Consequently, we present cancer specific exosome proteins and their unique roles in the interaction between cancer and its microenvironment. Clinically, these exosomes might provide useful biomarkers for cancer diagnosis and therapeutic tools for cancer treatment.
Unraveling proteome changes and potential regulatory proteins of bovine follicular Granulosa cells by mass spectrometry and multi-omics analysis.
Hou S, Hao Q, Zhu Z … +4 more , Xu D, Liu W, Lyu L, Li P
Proteome Sci · 2019 · PMID 31673248 · Full text
BACKGROUND: In previous study, we performed next-gene sequencing to investigate the differentially expressed transcripts of bovine follicular granulosa cells (GCs) at dominant follicle (DF) and subordinate follicle (SF)...
BACKGROUND: In previous study, we performed next-gene sequencing to investigate the differentially expressed transcripts of bovine follicular granulosa cells (GCs) at dominant follicle (DF) and subordinate follicle (SF) stages during first follicular wave. Present study is designed to further identify the key regulatory proteins and signaling pathways associated with follicular development using label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) and multi-omics data analysis approach. METHODS: DF and SF from three cattle were collected by daily ultrasonography. The GCs were isolated from each follicle, total proteins were digested by trypsin, and then proteomic analyzed via LC-MS/MS, respectively. Proteins identified were retrieved from Uniprot-COW fasta database, and differentially expressed proteins were used to functional enrichment and KEGG pathway analysis. Proteome data and transcriptome data obtained from previous studies were integrated. RESULTS: Total 3409 proteins were identified from 30,321 peptides (FDR ≤0.01) obtained from LC-MS/MS analysis and 259 of them were found to be differentially expressed at different stage of follicular development (fold Change > 2, < 0.05). KEGG pathway analysis of proteome data revealed important signaling pathways associated with follicular development, multi-omics data analysis results showed 13 proteins were identified as being differentially expressed in DF versus SF. CONCLUSIONS: This study represents the first investigation of transcriptome and proteome of bovine follicles and offers essential information for future investigation of DF and SF in cattle. It also will enrich the theory of animal follicular development.
Proteomic analysis of human periodontal ligament cells under hypoxia.
Li Q, Luo T, Lu W … +3 more , Yi X, Zhao Z, Liu J
Proteome Sci · 2019 · PMID 31496921 · Full text
BACKGROUND: The periodontal ligament is essential for homeostasis of periodontal tissue. A hypoxic milieu of the periodontal tissue is generated under periodontitis or during orthodontic treatment, which affects the peri...
BACKGROUND: The periodontal ligament is essential for homeostasis of periodontal tissue. A hypoxic milieu of the periodontal tissue is generated under periodontitis or during orthodontic treatment, which affects the periodontal and bone remodelling process. Here, we provide a comprehensive proteomic characterization of periodontal ligament cells under hypoxic conditions, aiming to reveal previously unappreciated biological changes and to help advance hypoxia-based therapeutic strategies for periodontal diseases. METHODS: Human periodontal ligament cells (hPDLCs) were characterized using immunohistochemistry (IHC) and flow cytometry (FACS). Successful hypoxia treatment of hPDLCs with 1% O was confirmed by qRT-PCR. Proliferation was evaluated using an MTT assay. The proteomic expression profile under hypoxia was studied with the isobaric tags for relative and absolute quantification (iTRAQ) approach followed by protein identification and bioinformatic analysis, and western blot verification was performed. RESULTS: The hPDLCs were positive for vimentin, CD73 and CD105 and negative for keratin, CD34 and CD45. After hypoxia treatment, the mRNA expression of hypoxia-inducible factor 1a ( was upregulated. The proliferation rate was elevated during the first 6 h but decreased from 6 h to 72 h. A total of 220 differentially expressed proteins were quantified in hPDLCs under hypoxia (1% O, 24 h), including 153 upregulated and 67 downregulated proteins, five of which were verified by western blot analysis. The Gene Ontology enriched terms included the energy metabolic process, membrane-bound organelle and vesicle, and protein binding terms. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated several involved pathways, including glycolysis/gluconeogenesis, biosynthesis of amino acids, the HIF-1 signalling pathway, and focal adhesion. A protein-protein interaction (PPI) network demonstrated the dominant role of autophagy over apoptosis under hypoxia. CONCLUSION: The proteomic profile of hPDLCs under hypoxia was mainly related to energy metabolism, autophagy, and responses to stimuli such as adhesion and inflammation. Previously unrecognized proteins including solute carrier family proteins, heat shock proteins, ubiquitination-related enzymes, collagen and S100 family proteins are involved in adaptive response to hypoxia in hPDLCs and are thus of great research interest in future work.
Comparison of enzymatic activities and proteomic profiles of grown on different carbon sources.
Sechovcová H, Kulhavá L, Fliegerová K … +4 more , Trundová M, Morais D, Mrázek J, Kopečný J
Proteome Sci · 2019 · PMID 31168299 · Full text
BACKGROUND: The rumen microbiota is one of the most complex consortia of anaerobes, involving archaea, bacteria, protozoa, fungi and phages. They are very effective at utilizing plant polysaccharides, especially cellulos...
BACKGROUND: The rumen microbiota is one of the most complex consortia of anaerobes, involving archaea, bacteria, protozoa, fungi and phages. They are very effective at utilizing plant polysaccharides, especially cellulose and hemicelluloses. The most important hemicellulose decomposers are clustered with the genus . As the related species differ in their range of hydrolytic activities and substrate preferences, was selected as one of the most effective isolates and thus suitable for proteomic studies on substrate comparisons in the extracellular fraction. The genome is the biggest in the butyrivibria cluster and is focused on "environmental information processing" and "carbohydrate metabolism". METHODS: The study of the effect of carbon source on 3071 was based on cultures grown on four substrates: xylose, glucose, xylan, xylan with 25% glucose. The enzymatic activities were studied by spectrophotometric and zymogram methods. Proteomic study was based on genomics, 2D electrophoresis and nLC/MS (Bruker Daltonics) analysis. RESULTS: Extracellular β-endoxylanase as well as xylan β-xylosidase activities were induced with xylan. The presence of the xylan polymer induced hemicellulolytic enzymes and increased the protein fraction in the interval from 40 to 80 kDa. 2D electrophoresis with nLC/MS analysis of extracellular 3071 proteins found 14 diverse proteins with significantly different expression on the tested substrates. CONCLUSION: The comparison of four carbon sources resulted in the main significant changes in proteome occurring outside the fibrolytic cluster of proteins. The affected proteins mainly belonged to the glycolysis and protein synthesis cluster.
Hepatic protein Carbonylation profiles induced by lipid accumulation and oxidative stress for investigating cellular response to non-alcoholic fatty liver disease in vitro.
Chienwichai P, Reamtong O, Boonyuen U … +5 more , Pisitkun T, Somparn P, Tharnpoophasiam P, Worakhunpiset S, Topanurak S
Proteome Sci · 2019 · PMID 30962768 · Full text
BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is caused by excessive accumulation of fat within the liver, leading to further severe conditions such as non-alcoholic steatohepatitis (NASH). Progression of healthy...
BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is caused by excessive accumulation of fat within the liver, leading to further severe conditions such as non-alcoholic steatohepatitis (NASH). Progression of healthy liver to steatosis and NASH is not yet fully understood in terms of process and response. Hepatic oxidative stress is believed to be one of the factors driving steatosis to NASH. Oxidative protein modification is the major cause of protein functional impairment in which alteration of key hepatic enzymes is likely to be a crucial factor for NAFLD biology. In the present study, we aimed to discover carbonylated protein profiles involving in NAFLD biology in vitro. METHODS: Hepatocyte cell line was used to induce steatosis with fatty acids (FA) in the presence and absence of menadione (oxidative stress inducer). Two-dimensional gel electrophoresis-based proteomics and dinitrophenyl hydrazine derivatization technique were used to identify carbonylated proteins. Sequentially, in order to view changes in protein carbonylation pathway, enrichment using Funrich algorithm was performed. The selected carbonylated proteins were validated with western blot and carbonylated sites were further identified by high-resolution LC-MS/MS. RESULTS: Proteomic results and pathway analysis revealed that carbonylated proteins are involved in NASH pathogenesis pathways in which most of them play important roles in energy metabolisms. Particularly, carbonylation level of ATP synthase subunit α (ATP5A), a key protein in cellular respiration, was reduced after FA and FA with oxidative stress treatment, whereas its expression was not altered. Carbonylated sites on this protein were identified and it was revealed that these sites are located in nucleotide binding region. Modification of these sites may, therefore, disturb ATP5A activity. As a consequence, the lower carbonylation level on ATP5A after FA treatment solely or with oxidative stress can increase ATP production. CONCLUSIONS: The reduction in carbonylated level of ATP5A might occur to generate more energy in response to pathological conditions, in our case, fat accumulation and oxidative stress in hepatocytes. This would imply the association between protein carbonylation and molecular response to development of steatosis and NASH.
Proteome modifications on tomato under extreme high light induced-stress.
Parrine D, Wu BS, Muhammad B … +4 more , Rivera K, Pappin D, Zhao X, Lefsrud M
Proteome Sci · 2018 · PMID 30534005 · Full text
BACKGROUND: Abiotic stress reduces photosynthetic yield and plant growth, negatively impacting global crop production and is a major constraint faced by agriculture. However, the knowledge on the impact on plants under e...
BACKGROUND: Abiotic stress reduces photosynthetic yield and plant growth, negatively impacting global crop production and is a major constraint faced by agriculture. However, the knowledge on the impact on plants under extremely high irradiance is limited. We present the first in-depth proteomics analysis of plants treated with a method developed by our research group to generate a light gradient using an extremely intense light. METHODS: The method consists of utilizing light emitting diodes (LED) to create a single spot at 24,000 μmol m s irradiance, generating three light stress levels. A light map and temperature profile were obtained during the light experiment. The proteins expressed in the treated tomato (, Heinz H1706) leaves were harvested 10 days after the treatment, allowing for the detection of proteins involved in a long-term recovery. A multiplex labeled proteomics method (iTRAQ) was analyzed by LC-MS/MS. RESULTS: A total of 3994 proteins were identified at 1% false discovery rate and matched additional quality filters. Hierarchical clustering analysis resulted in four types of patterns related to the protein expression, with one being directly linked to the increased LED irradiation. A total of 37 proteins were found unique to the least damaged leaf zone, while the medium damaged zone had 372 proteins, and the severely damaged presented unique 1003 proteins. Oxygen evolving complex and PSII complex proteins (PsbH, PsbS, PsbR and Psb28) were found to be abundant in the most damaged leaf zone. This leaf zone presented a protein involved in the salicylic acid response, while it was not abundant in the other leaf zones. The mRNA level of PsbR was significantly lower (1-fold) compared the control in the most damaged zone of the leaf, while Psb28 and PsbH were lower (1-fold) in the less damaged leaf zones. PsbS mRNA abundance in all leaf zones tested presented no statistically significant change from the control. CONCLUSIONS: We present the first characterization of the proteome changes caused by an extreme level of high-light intensity (24,000 μmol m s). The proteomics results show the presence of specific defense responses to each level of light intensity, with a possible involvement of proteins PsbH, Psb28, PsbR, and PsbS.
Key factors identified by proteomic analysis in maize ( L.) seedlings' response to long-term exposure to different phosphate levels.
Sun Y, Mu C, Liu X
Proteome Sci · 2018 · PMID 30479573 · Full text
BACKGROUND: Maize seedlings are constantly exposed to inorganic phosphate (Pi)-limited environments. To understand how maize cope with low Pi (LP) and high Pi (HP) conditions, physiological and global proteomic analysis...
BACKGROUND: Maize seedlings are constantly exposed to inorganic phosphate (Pi)-limited environments. To understand how maize cope with low Pi (LP) and high Pi (HP) conditions, physiological and global proteomic analysis of QXN233 genotype were performed under the long-term Pi starvation and supplementation. METHODS: We investigated the physiological response of QXN233 genotype to LP and HP conditions and detected the changes in ion fluxes by non-invasive micro-test technology and gene expression by quantitative real-time polymerase chain reaction. QXN233 was further assessed using vermiculite assay, and then proteins were isolated and identified by nano-liquid chromatography-mass spectrometry. RESULTS: A negative relationship was observed between Na and Pi, and Na efflux was enhanced under HP condition. Furthermore, a total of 681 and 1374 were identified in the leaves and roots, respectively, which were mostly involved in metabolism, ion transport, and stress response. Importantly, several key Pi transporters were identified for breeding potential. Several ion transporters demonstrated an elaborate interplay between Pi and other ions, together contributing to the growth of QXN233 seedlings. CONCLUSION: The results from this study provide insights into the response of maize seedlings to long-term Pi exposure.
Calcitonin gene-related peptide promotes proliferation and inhibits apoptosis in endothelial progenitor cells via inhibiting MAPK signaling.
Wu J, Liu S, Wang Z … +3 more , Ma S, Meng H, Hu J
Proteome Sci · 2018 · PMID 30473635 · Full text
BACKGROUND: Calcitonin gene-related peptide (CGRP) contributes to bone formation by stimulating bone marrow stromal cell (BMSC) proliferation and differentiation. However, the proliferative and apoptotic effects of CGRP...
BACKGROUND: Calcitonin gene-related peptide (CGRP) contributes to bone formation by stimulating bone marrow stromal cell (BMSC) proliferation and differentiation. However, the proliferative and apoptotic effects of CGRP on bone marrow-derived endothelial progenitor cells (EPCs) have not been investigated. METHODS: We tested the effects of CGRP on EPC proliferation and apoptosis by Cell Counting Kit-8, flow cytometry, and studied the effects of CGRP on the expression of proliferation- and apoptosis-related markers in EPCs and the underlying mitogen-activated protein kinase (MAPK) signalling pathway by quantitative polymerase chain reaction and western blotting. RESULTS: We detected EPC markers (CD34, CD133 and VEGFR-2) in 7-day cultures and found that CGRP (10-10 M) promoted the proliferation of cultured EPCs, with a peak increase of 30% at 10 M CGRP. CGRP also upregulated the expression of proliferation-associated genes, including cyclin D1 and cyclin E, and increased the percentages of G2/M-phase and S-phase cells after incubation 72 h. CGRP inhibited serum deprivation (SD)-induced apoptosis in EPCs after 24 and 48 h and downregulated the expression of apoptosis-related genes, including caspase-3, caspase-8, caspase-9 and Bax. Phosphorylated (p-)ERK1/2, p-p38 and p-JNK protein levels in EPCs treated with CGRP were significantly lower than those in untreated EPCs. Pre-treatment with the calcitonin receptor-like receptor (CRLR) antagonist CGRP8-37 or a MAPK pathway inhibitor (PD98059, SB203580 or SP600125) completely or partially reversed the pro-proliferative and anti-apoptotic effects and the reduced p-ERK1/2, p-p38 and p-JNK expression induced by CGRP. CONCLUSION: Our results show that CGRP exerts pro-proliferative and anti-apoptotic effects on EPCs and may act by inhibiting MAPK pathways.
Comparative proteomic analysis of response to high temperature stress.
Fan M, Sun X, Liao Z … +3 more , Wang J, Li Y, Xu N
Proteome Sci · 2018 · PMID 30386183 · Full text
BACKGROUND: belongs to green macroalgae and is the dominant species of green tide. It is distributed worldwide and is therefore subject to high-temperature stress during the growth process. However, the adaptation mecha...
BACKGROUND: belongs to green macroalgae and is the dominant species of green tide. It is distributed worldwide and is therefore subject to high-temperature stress during the growth process. However, the adaptation mechanisms of the response of to high temperatures have not been clearly investigated yet. METHODS: In this study, isobaric tags for relative and absolute quantitation (iTRAQ) labelling was applied in combination with the liquid chromatography-tandem mass spectrometry (LC-MS/MS) to conduct comparative proteomic analysis of the response of to high-temperature stress and to elucidate the involvement of this response in adaptation mechanisms. Differentially expressed proteins (DEPs) of under high temperature (denote UpHT) compared with the control (UpC) were identified. Bioinformatic analyses including GO analysis, pathway analysis, and pathway enrichment analysis was performed to analyse the key metabolic pathways that underlie the thermal tolerance mechanism through protein networks. Quantitative real-time PCR and western blot were performed to validate selected proteins. RESULTS: In the present study, 1223 DEPs were identified under high temperature compared with the control, which included 790 up-regulated and 433 down-regulated proteins. The high-temperature stimulus mainly induced the expression of glutathione S-transferase, heat shock protein, ascorbate peroxidase, manganese superoxide dismutase, ubiquitin-related protein, lhcSR, rubisco activase, serine/threonine protein kinase 2, adenylate kinase, Ca-dependent protein kinase (CDPK), disease resistance protein EDS1, metacaspase type II, NDPK2a, 26S proteasome regulatory subunit, ubiquinone oxidoreductase, ATP synthase subunit, SnRK2s, and cytochrome P450. The down-regulated proteins were photosynthesis-related proteins, glutathione reductase, catalase-peroxidase, thioredoxin, thioredoxin peroxidase, PP2C, and carbon fixation-related proteins. Furthermore, biological index analysis indicated that protein content and SOD activity decreased; the value of Fv/Fm dropped to the lowest point after culture for 96 h. However, APX activity and MDA content increased under high temperature. CONCLUSION: The present study implied an increase in proteins that were associated with the stress response, oxidative phosphorylation, the cytokinin signal transduction pathway, the abscisic acid signal transduction pathway, and the glutathione metabolism pathway. Proteins that were associated with photosynthesis, carbon fixation in photosynthesis organisms, and the photosynthesis antenna protein pathway were decreased. These pathways played a pivotal role in high temperature regulation. These novel proteins provide a good starting point for further research into their functions using genetic or other approaches. These findings significantly improve the understanding of the molecular mechanisms involved in the tolerance of algae to high-temperature stress.
Global phosphoproteomic analysis identifies SRMS-regulated secondary signaling intermediates.
Goel RK, Meyer M, Paczkowska M … +5 more , Reimand J, Vizeacoumar F, Vizeacoumar F, Lam TT, Lukong KE
Proteome Sci · 2018 · PMID 30140170 · Full text
BACKGROUND: The non-receptor tyrosine kinase, SRMS (Src-related kinase lacking C-terminal regulatory tyrosine and N-terminal myristoylation sites) is a member of the BRK family kinases (BFKs) which represents an evolutio...
BACKGROUND: The non-receptor tyrosine kinase, SRMS (Src-related kinase lacking C-terminal regulatory tyrosine and N-terminal myristoylation sites) is a member of the BRK family kinases (BFKs) which represents an evolutionarily conserved relative of the Src family kinases (SFKs). Tyrosine kinases are known to regulate a number of cellular processes and pathways via phosphorylating substrate proteins directly and/or by partaking in signaling cross-talks leading to the indirect modulation of various signaling intermediates. In a previous study, we profiled the tyrosine-phosphoproteome of SRMS and identified multiple candidate substrates of the kinase. The broader cellular signaling intermediates of SRMS are unknown. METHODS: In order to uncover the broader SRMS-regulated phosphoproteome and identify the SRMS-regulated indirect signaling intermediates, we performed label-free global phosphoproteomics analysis on cells expressing wild-type SRMS. Using computational database searching and bioinformatics analyses we characterized the dataset. RESULTS: Our analyses identified 60 hyperphosphorylated (phosphoserine/phosphothreonine) proteins mapped from 140 hyperphosphorylated peptides. Bioinfomatics analyses identified a number of significantly enriched biological and cellular processes among which DNA repair pathways were found to be upregulated while apoptotic pathways were found to be downregulated. Analyses of motifs derived from the upregulated phosphosites identified Casein kinase 2 alpha (CK2α) as one of the major potential kinases contributing to the SRMS-dependent indirect regulation of signaling intermediates. CONCLUSIONS: Overall, our phosphoproteomics analyses identified serine/threonine phosphorylation dynamics as important secondary events of the SRMS-regulated phosphoproteome with implications in the regulation of cellular and biological processes.
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