Monitoring of host cell proteins(HCPs)during the manufacturing of monoclonal antibodies(mAb)has become a critical requirement to provide effective and safe drug products.Enzyme-linked immunosorbent assays are still th...Monitoring of host cell proteins(HCPs)during the manufacturing of monoclonal antibodies(mAb)has become a critical requirement to provide effective and safe drug products.Enzyme-linked immunosorbent assays are still the gold standard methods for the quantification of protein impurities.However,this technique has several limitations and does,among others,not enable the precise identification of proteins.In this context,mass spectrometry(MS)became an alternative and orthogonal method that delivers qualitative and quantitative information on all identified HCPs.However,in order to be routinely implemented in biopharmaceutical companies,liquid chromatography-MS based methods still need to be standardized to provide highest sensitivity and robust and accurate quantification.Here,we present a promising MS-based analytical workflow coupling the use of an innovative quantification standard,the HCP Profiler solution,with a spectral library-based data-independent acquisition(DIA)method and strict data validation criteria.The performances of the HCP Profiler solution were compared to more conventional standard protein spikes and the DIA approach was benchmarked against a classical datadependent acquisition on a series of samples produced at various stages of the manufacturing process.While we also explored spectral library-free DIA interpretation,the spectral library-based approach still showed highest accuracy and reproducibility(coefficients of variation<10%)with a sensitivity down to the sub-ng/mg mAb level.Thus,this workflow is today mature to be used as a robust and straightforward method to support mAb manufacturing process developments and drug products quality control.展开更多
Ensuring the removal of host cell proteins(HCPs) during downstream processing of recombinant proteins such as monoclonal antibodies(m Abs) remains a challenge.Since residual HCPs might affect product stability or safe...Ensuring the removal of host cell proteins(HCPs) during downstream processing of recombinant proteins such as monoclonal antibodies(m Abs) remains a challenge.Since residual HCPs might affect product stability or safety,constant monitoring is required to demonstrate their removal to be below the regulatory accepted level of 100 ng/mg.The current standard analytical approach for this procedure is based on ELISA;however,this approach only measures the overall HCP content.Therefore,the use of orthogonal methods,such as liquid chromatography-mass spectrometry(LC-MS),has been established,as it facilitates the quantitation of total HCPs as well as the identification and quantitation of the individual HCPs present.In the present study,a workflow for HCP detection and quantitation using an automated magnetic bead-based sample preparation,in combination with a data-independent acquisition(DIA) LC-MS analysis,was established.Employing the same instrumental setup commonly used for peptide mapping analysis of m Abs allows for its quick and easy implementation into pre-existing workflows,avoiding the need for dedicated instrumentation or personnel.Thereby,quantitation of HCPs over a broad dynamic range was enabled to allow monitoring of problematic HCPs or to track changes upon altered bioprocessing conditions.展开更多
Background In poultry, the smooth transition of follicles from the preovulatory-to-postovulatory phase impacts egg production in hens and can benefit the poultry industry. However, the regulatory mechanism underlying ...Background In poultry, the smooth transition of follicles from the preovulatory-to-postovulatory phase impacts egg production in hens and can benefit the poultry industry. However, the regulatory mechanism underlying follicular ovulation in avians is a complex biological process that remains unclear.Results Critical biochemical events involved in ovulation in domestic chickens(Gallus gallus) were evaluated by transcriptomics, proteomics, and in vitro assays. Comparative transcriptome analyses of the largest preovulatory follicle(F1) and postovulatory follicle(POF1) in continuous laying(CL) and intermittent laying(IL) chickens indicated the greatest difference between CL_F1 and IL_F1, with 950 differentially expressed genes(DEGs), and the smallest difference between CL_POF1 and IL_POF1, with 14 DEGs. Additionally, data-independent acquisition proteomics revealed 252 differentially abundant proteins between CL_F1 and IL_F1. Perivitelline membrane synthesis, steroid biosynthesis, lysosomes, and oxidative phosphorylation were identified as pivotal pathways contributing to ovulation regulation. In particular, the regulation of zona pellucida sperm-binding protein 3, plasminogen activator, cathepsin A, and lactate dehydrogenase A(LDHA) was shown to be essential for ovulation. Furthermore, the inhibition of LDHA decreased cell viability and promoted apoptosis of ovarian follicles in vitro.Conclusions This study reveals several important biochemical events involved in the process of ovulation, as well as crucial role of LDHA. These findings improve our understanding of ovulation and its regulatory mechanisms in avian species.展开更多
The milk fat globule membrane(MFGM)is a complex structure with numerous functions,and its composition is affected by many factors.There have been few systematic investigations on goat MFGM proteome profiling during la...The milk fat globule membrane(MFGM)is a complex structure with numerous functions,and its composition is affected by many factors.There have been few systematic investigations on goat MFGM proteome profiling during lactation.Individual milk samples from 15 healthy dairy goats were obtained at six lactation time points for investigation of the MFGM proteome using both data-independent acquisition(DIA)and data-dependent acquisition(DDA)proteomics techniques combined with multivariate statistical analysis.Using the DIA method,890 variably abundant MFGM proteins were discovered throughout the lactation cycle.From 1 to 240 d,butyrophilin subfamily 1 member A1,lipoprotein lipase,perilipin-2,and adipose triglyceride lipase were upregulated,while APOE,complement C3,clusterin,and IgG were downregulated.Furthermore,from 1 to 90 d,annexin A1,annexin A2,and antithrombin-ll were downregulated,then upregulated by d 240.Albumin had a high degree of connectedness,indicating that it was a key protein,according to protein-protein interaction research.Overall,our findings gave new insights into the biological features of MFGM protein in goat milk throughout lactation,which may aid in the creation of specialized MFGM products and infant formula.展开更多
Ogura cytoplasmic male sterility(Ogura CMS)is extensively applied in hybrid seed production in cruciferous crops.However,the posttranscriptional molecular basis of Ogura CMS in cruciferous crops remains elusive.Here,a...Ogura cytoplasmic male sterility(Ogura CMS)is extensively applied in hybrid seed production in cruciferous crops.However,the posttranscriptional molecular basis of Ogura CMS in cruciferous crops remains elusive.Here,a data-independent acquisition-based proteomic approach coupled with a parallel reaction monitoring-based targeted proteomic assay was used to analyze the proteome dynamics of Ogura CMS cabbage line RM and its maintainer line RF during floral bud development to obtain insights into the mechanism underlying Ogura CMS in cruciferous crops.A total of 9162 proteins corresponding to 61464 peptides were identified in RM and RF floral buds.The proteomic fluctuation of RM was weaker than that of RF.Differences in protein expression between RM and RF gradually enlarged with floral bud development.Fifteen continually up-regulated and eight continually down-regulated proteins were found in RM relative to RF throughout floral bud development.Differentially expressed proteins between RM and RF during floral bud development were implicated in the endoplasmic reticulum(ER)-associated protein processing pathway,in which most of them exhibited down-regulated expression in RM.These data suggest that ER-associated protein processing may be involved in pollen abortion in Ogura CMS cabbage by inhibiting the expression of critical factors.Our findings not only deepen the understanding of the molecular mechanisms of Ogura CMS in cruciferous crops but also provide better guidance for applying Ogura CMS in the hybrid breeding of cruciferous crops.展开更多
BACKGROUND Colorectal cancer(CRC)is the second leading cause of cancer-related death,with high morbidity worldwide.There is an urgent need to find reliable diagnostic biomarkers of CRC and explore the underlying molec...BACKGROUND Colorectal cancer(CRC)is the second leading cause of cancer-related death,with high morbidity worldwide.There is an urgent need to find reliable diagnostic biomarkers of CRC and explore the underlying molecular mechanisms.Exosomes are involved in intercellular communication and participate in multiple pathological processes,serving as an important part of the tumor microenvironment.AIM To investigate the proteomic characteristics of CRC tumor-derived exosomes and to identify candidate exosomal protein markers for CRC.METHODS In this study,10 patients over 50 years old who were diagnosed with moderately differentiated adenocarcinoma were recruited.We paired CRC tissues and adjacent normal intestinal tissues(>5 cm)to form the experimental and control groups.Purified exosomes were extracted separately from each tissue sample.Data-independent acquisition mass spectrometry was implemented in 8 matched samples of exosomes to explore the proteomic expression profiles,and differentially expressed proteins(DEPs)were screened by bioinformatics analysis.Promising exosomal proteins were verified using parallel reaction monitoring(PRM)analysis in 10 matched exosome samples.RESULTS A total of 1393 proteins were identified in the CRC tissue group,1304 proteins were identified in the adjacent tissue group,and 283 proteins were significantly differentially expressed between them.Enrichment analysis revealed that DEPs were involved in multiple biological processes related to cytoskeleton construction,cell movement and migration,immune response,tumor growth and telomere metabolism,as well as ECM-receptor interaction,focal adhesion and mTOR signaling pathways.Six differentially expressed exosomal proteins(NHP2,OLFM4,TOP1,SAMP,TAGL and TRIM28)were validated by PRM analysis and evaluated by receiver operating characteristic curve(ROC)analysis.The area under the ROC curve was 0.93,0.96,0.97,0.78,0.75,and 0.88(P<0.05)for NHP2,OLFM4,TOP1,SAMP,TAGL,and TRIM28,respectively,indicating their good ability to distinguish CRC tissues from adjacent intestinal tissues.CONCLUSION In our study,comprehensive proteomic profiles were obtained for CRC tissue exosomes.Six exosomal proteins,NHP2,OLFM4,TOP1,SAMP,TAGL and TRIM28,may be promising diagnostic markers and effective therapeutic targets for CRC,but further experimental investigation is needed.展开更多
The effects of putrescine on improving rice growth under aluminum(Al)toxicity conditions have been previously demonstrated,however,the underlying mechanism remains unclear.In this study,treatment with 50 pmol/L Al sig...The effects of putrescine on improving rice growth under aluminum(Al)toxicity conditions have been previously demonstrated,however,the underlying mechanism remains unclear.In this study,treatment with 50 pmol/L Al significantly decreased rice root growth and whole rice dry weight,inhibited Ca2+uptake,decreased ATP syn thesis,and in creased Al,H2O2 and malon dialdehyde(MDA)con tents,whereas the application of putrescine mitigated these negative effects.Putrescine increased root growth and total dry weight of rice,reduced total Al content,decreased H2O2 and MDA contents by increasing antioxidant enzyme(superoxide dismutase,peroxidase,catalase and glutathione S・transferase)activities,increased Ca2+uptake and energy product!oru Proteomic analyses using data-independent acquisition successfully identified 7934 proteins,and 59 representative proteins exhibiting fold-change values higher than 1.5 were randomly selected.From the results of the proteomic and biochemical analyses,we found that putrescine significantly inhibited ethylene biosynthesis and phosphorus uptake in rice roots,increased pectin methylation,decreased pectin content and apoplastic Al deposit!on in rice roots.Putrescine also alleviated Al toxicity by repairing damaged DNA and increasing the proteins involved in maintaining plasma membra ne integrity and normal cell proliferation.These fin dings improve our understanding of how putrescine affects the rice response to Al toxicity,which will facilitate further studies on environmental protection,crop safety,in novations in rice performance and real-world producti on.展开更多
Lipidomics coverage improvement is essential for functional lipid and pathway construction.A powerful approach to discovering organism lipidome is to combine various data acquisitions,such as full scan mass spectromet...Lipidomics coverage improvement is essential for functional lipid and pathway construction.A powerful approach to discovering organism lipidome is to combine various data acquisitions,such as full scan mass spectrometry(full MS),data-dependent acquisition(DDA),and data-independent acquisition(DIA).Caenorhabditis elegans(C.elegans)is a useful model for discovering toxic-induced metabolism,highthroughput drug screening,and a variety of human disease pathways.To determine the lipidome of C.elegans and investigate lipid disruption from the molecular level to the system biology level,we used integrative data acquisition.The methyl-tert-butyl ether method was used to extract L4 stage C.elegans after exposure to triclosan(TCS),perfluorooctanoic acid,and nanopolystyrene(nPS).Full MS,DDA,and DIA integrations were performed to comprehensively profile the C.elegans lipidome by Q-Exactive Plus MS.All annotated lipids were then analyzed using lipid ontology and pathway analysis.We annotated up to 940 lipids from 20 lipid classes involved in various functions and pathways.The biological investigations revealed that when C.elegans were exposed to nPS,lipid droplets were disrupted,whereas plasma membrane-functionalized lipids were likely to be changed in the TCS treatment group.The nPS treatment caused a significant disruption in lipid storage.Triacylglycerol,glycerophospholipid,and ether class lipids were those primarily hindered by toxicants.Finally,toxicant exposure frequently involved numerous lipid-related pathways,including the phosphoinositide 3-kinase/protein kinase B pathway.In conclusion,an integrative data acquisition strategy was used to characterize the C.elegans lipidome,providing valuable biological insights into hypothesis generation and validation.展开更多
Lung cancer produces a high incidence of malignant tumors.There have been many studies on lung cancer using mass spectrometry(MS)technologies.However,most studies have focused on humoral samples.In this work,26 pairs ...Lung cancer produces a high incidence of malignant tumors.There have been many studies on lung cancer using mass spectrometry(MS)technologies.However,most studies have focused on humoral samples.In this work,26 pairs of tissue samples(tumor vs.para-tumor)from patients with lung cancer were analyzed using liquid chromatography-tandem MS(LC-MS/MS)with data-independent acquisition mode.In total,3152 proteins were quantified from tissue samples with high confidence,including 189 up-regulated and 522 down-regulated proteins(tumor vs.para-tumor).In addition,79 and 690 proteins were identified only in para-cancerous samples and cancerous samples,respectively.The results from bio-informatics tools indicated that altered proteins like PEBP1,HRG and LYZ could be ideal reservoirs for screening the potential biomarkers for lung cancer.It is believed these tissue-specific proteomics results will assist in the studies of lung cancer.展开更多
To address the increasing need for detecting and validating protein biomarkers in clinical specimens,mass spectrometry(MS)-based targeted proteomic techniques,including the selected reaction monitoring(SRM),parallel r...To address the increasing need for detecting and validating protein biomarkers in clinical specimens,mass spectrometry(MS)-based targeted proteomic techniques,including the selected reaction monitoring(SRM),parallel reaction monitoring(PRM),and massively parallel dataindependent acquisition(DIA),have been developed.For optimal performance,they require the fragment ion spectra of targeted peptides as prior knowledge.In this report,we describe a MS pipeline and spectral resource to support targeted proteomics studies for human tissue samples.To build the spectral resource,we integrated common open-source MS computational tools to assemble a freely accessible computational workflow based on Docker.We then applied the workflow to generate DPHL,a comprehensive DIA pan-human library,from 1096 data-dependent acquisition(DDA)MS raw files for 16 types of cancer samples.This extensive spectral resource was then applied to a proteomic study of 17 prostate cancer(PCa)patients.Thereafter,PRM validation was applied to a larger study of 57 PCa patients and the differential expression of three proteins in prostate tumor was validated.As a second application,the DPHL spectral resource was applied to a study consisting of plasma samples from 19 diffuse large B cell lymphoma(DLBCL)patients and 18 healthy control subjects.Differentially expressed proteins between DLBCL patients and healthy control subjects were detected by DIA-MS and confirmed by PRM.These data demonstrate that the DPHL supports DIA and PRM MS pipelines for robust protein biomarker discovery.DPHL is freely accessible at https://www.iprox.org/page/project.html?id=IPX0001400000.展开更多
基金supported by the“Association Nationale de la Recherche et de la Technologie”and UCB Pharma S.A.(Belgium and France)via the CIFRE fellowship of Steve Hessmannsupported by the“Agence Nationale de la Recherche”via the French Proteomic Infrastructure ProFI FR2048(ANR-10-INBS-08-03).
文摘Monitoring of host cell proteins(HCPs)during the manufacturing of monoclonal antibodies(mAb)has become a critical requirement to provide effective and safe drug products.Enzyme-linked immunosorbent assays are still the gold standard methods for the quantification of protein impurities.However,this technique has several limitations and does,among others,not enable the precise identification of proteins.In this context,mass spectrometry(MS)became an alternative and orthogonal method that delivers qualitative and quantitative information on all identified HCPs.However,in order to be routinely implemented in biopharmaceutical companies,liquid chromatography-MS based methods still need to be standardized to provide highest sensitivity and robust and accurate quantification.Here,we present a promising MS-based analytical workflow coupling the use of an innovative quantification standard,the HCP Profiler solution,with a spectral library-based data-independent acquisition(DIA)method and strict data validation criteria.The performances of the HCP Profiler solution were compared to more conventional standard protein spikes and the DIA approach was benchmarked against a classical datadependent acquisition on a series of samples produced at various stages of the manufacturing process.While we also explored spectral library-free DIA interpretation,the spectral library-based approach still showed highest accuracy and reproducibility(coefficients of variation<10%)with a sensitivity down to the sub-ng/mg mAb level.Thus,this workflow is today mature to be used as a robust and straightforward method to support mAb manufacturing process developments and drug products quality control.
基金funding from Thermo Fisher Scientific as part of a funded collaborative agreement with NIBR。
文摘Ensuring the removal of host cell proteins(HCPs) during downstream processing of recombinant proteins such as monoclonal antibodies(m Abs) remains a challenge.Since residual HCPs might affect product stability or safety,constant monitoring is required to demonstrate their removal to be below the regulatory accepted level of 100 ng/mg.The current standard analytical approach for this procedure is based on ELISA;however,this approach only measures the overall HCP content.Therefore,the use of orthogonal methods,such as liquid chromatography-mass spectrometry(LC-MS),has been established,as it facilitates the quantitation of total HCPs as well as the identification and quantitation of the individual HCPs present.In the present study,a workflow for HCP detection and quantitation using an automated magnetic bead-based sample preparation,in combination with a data-independent acquisition(DIA) LC-MS analysis,was established.Employing the same instrumental setup commonly used for peptide mapping analysis of m Abs allows for its quick and easy implementation into pre-existing workflows,avoiding the need for dedicated instrumentation or personnel.Thereby,quantitation of HCPs over a broad dynamic range was enabled to allow monitoring of problematic HCPs or to track changes upon altered bioprocessing conditions.
基金supported by the National Key Research and Development Program of China (2022YFD1600902)Key Research and Development Program of Shandong (2022LZGC013)China Agriculture Research System (CARS-40)。
文摘Background In poultry, the smooth transition of follicles from the preovulatory-to-postovulatory phase impacts egg production in hens and can benefit the poultry industry. However, the regulatory mechanism underlying follicular ovulation in avians is a complex biological process that remains unclear.Results Critical biochemical events involved in ovulation in domestic chickens(Gallus gallus) were evaluated by transcriptomics, proteomics, and in vitro assays. Comparative transcriptome analyses of the largest preovulatory follicle(F1) and postovulatory follicle(POF1) in continuous laying(CL) and intermittent laying(IL) chickens indicated the greatest difference between CL_F1 and IL_F1, with 950 differentially expressed genes(DEGs), and the smallest difference between CL_POF1 and IL_POF1, with 14 DEGs. Additionally, data-independent acquisition proteomics revealed 252 differentially abundant proteins between CL_F1 and IL_F1. Perivitelline membrane synthesis, steroid biosynthesis, lysosomes, and oxidative phosphorylation were identified as pivotal pathways contributing to ovulation regulation. In particular, the regulation of zona pellucida sperm-binding protein 3, plasminogen activator, cathepsin A, and lactate dehydrogenase A(LDHA) was shown to be essential for ovulation. Furthermore, the inhibition of LDHA decreased cell viability and promoted apoptosis of ovarian follicles in vitro.Conclusions This study reveals several important biochemical events involved in the process of ovulation, as well as crucial role of LDHA. These findings improve our understanding of ovulation and its regulatory mechanisms in avian species.
基金This work was supportedby theNational KeyR&D Program of China(2022YFD1301005)the Shandong Provincial Natural Science Foundation,China(ZR2022MC184)the High-level Talents Foundation of Qingdao Agricultural University,China(665/1120053,665/1120080).
文摘The milk fat globule membrane(MFGM)is a complex structure with numerous functions,and its composition is affected by many factors.There have been few systematic investigations on goat MFGM proteome profiling during lactation.Individual milk samples from 15 healthy dairy goats were obtained at six lactation time points for investigation of the MFGM proteome using both data-independent acquisition(DIA)and data-dependent acquisition(DDA)proteomics techniques combined with multivariate statistical analysis.Using the DIA method,890 variably abundant MFGM proteins were discovered throughout the lactation cycle.From 1 to 240 d,butyrophilin subfamily 1 member A1,lipoprotein lipase,perilipin-2,and adipose triglyceride lipase were upregulated,while APOE,complement C3,clusterin,and IgG were downregulated.Furthermore,from 1 to 90 d,annexin A1,annexin A2,and antithrombin-ll were downregulated,then upregulated by d 240.Albumin had a high degree of connectedness,indicating that it was a key protein,according to protein-protein interaction research.Overall,our findings gave new insights into the biological features of MFGM protein in goat milk throughout lactation,which may aid in the creation of specialized MFGM products and infant formula.
基金supported by Heilongjiang Provincial Natural Science Foundation of China(Grant No.YQ2022C012)China Postdoctoral Science Foundation(Grant No.2022MD713728)+1 种基金Heilongjiang Provincial Postdoctoral Fund(Grant No.LBHZ21046)the Open Project of Key Laboratory of Biology and Genetic Improvement of Horticultural Crops(Northeast Region),Ministry of Agriculture and Rural Affairs,and National Key Research and Development Program of China(Grant No.2023YFD1201501).
文摘Ogura cytoplasmic male sterility(Ogura CMS)is extensively applied in hybrid seed production in cruciferous crops.However,the posttranscriptional molecular basis of Ogura CMS in cruciferous crops remains elusive.Here,a data-independent acquisition-based proteomic approach coupled with a parallel reaction monitoring-based targeted proteomic assay was used to analyze the proteome dynamics of Ogura CMS cabbage line RM and its maintainer line RF during floral bud development to obtain insights into the mechanism underlying Ogura CMS in cruciferous crops.A total of 9162 proteins corresponding to 61464 peptides were identified in RM and RF floral buds.The proteomic fluctuation of RM was weaker than that of RF.Differences in protein expression between RM and RF gradually enlarged with floral bud development.Fifteen continually up-regulated and eight continually down-regulated proteins were found in RM relative to RF throughout floral bud development.Differentially expressed proteins between RM and RF during floral bud development were implicated in the endoplasmic reticulum(ER)-associated protein processing pathway,in which most of them exhibited down-regulated expression in RM.These data suggest that ER-associated protein processing may be involved in pollen abortion in Ogura CMS cabbage by inhibiting the expression of critical factors.Our findings not only deepen the understanding of the molecular mechanisms of Ogura CMS in cruciferous crops but also provide better guidance for applying Ogura CMS in the hybrid breeding of cruciferous crops.
基金Supported by National Key Development Plan for Precision Medicine Research,No.2017YFC0910002。
文摘BACKGROUND Colorectal cancer(CRC)is the second leading cause of cancer-related death,with high morbidity worldwide.There is an urgent need to find reliable diagnostic biomarkers of CRC and explore the underlying molecular mechanisms.Exosomes are involved in intercellular communication and participate in multiple pathological processes,serving as an important part of the tumor microenvironment.AIM To investigate the proteomic characteristics of CRC tumor-derived exosomes and to identify candidate exosomal protein markers for CRC.METHODS In this study,10 patients over 50 years old who were diagnosed with moderately differentiated adenocarcinoma were recruited.We paired CRC tissues and adjacent normal intestinal tissues(>5 cm)to form the experimental and control groups.Purified exosomes were extracted separately from each tissue sample.Data-independent acquisition mass spectrometry was implemented in 8 matched samples of exosomes to explore the proteomic expression profiles,and differentially expressed proteins(DEPs)were screened by bioinformatics analysis.Promising exosomal proteins were verified using parallel reaction monitoring(PRM)analysis in 10 matched exosome samples.RESULTS A total of 1393 proteins were identified in the CRC tissue group,1304 proteins were identified in the adjacent tissue group,and 283 proteins were significantly differentially expressed between them.Enrichment analysis revealed that DEPs were involved in multiple biological processes related to cytoskeleton construction,cell movement and migration,immune response,tumor growth and telomere metabolism,as well as ECM-receptor interaction,focal adhesion and mTOR signaling pathways.Six differentially expressed exosomal proteins(NHP2,OLFM4,TOP1,SAMP,TAGL and TRIM28)were validated by PRM analysis and evaluated by receiver operating characteristic curve(ROC)analysis.The area under the ROC curve was 0.93,0.96,0.97,0.78,0.75,and 0.88(P<0.05)for NHP2,OLFM4,TOP1,SAMP,TAGL,and TRIM28,respectively,indicating their good ability to distinguish CRC tissues from adjacent intestinal tissues.CONCLUSION In our study,comprehensive proteomic profiles were obtained for CRC tissue exosomes.Six exosomal proteins,NHP2,OLFM4,TOP1,SAMP,TAGL and TRIM28,may be promising diagnostic markers and effective therapeutic targets for CRC,but further experimental investigation is needed.
基金supported by the Zhejiang Provincial Natural Science Foundation of China(Grant No.LQ19C020007)the National Natural Science Foundation of China(Grant Nos.31901452,31771733,32001104 and 31872857)+2 种基金the Key Research and Development Program of Zhejiang Province,China(Grant No.2021C02002)the Open Project of State Key Laboratory of Rice Biology of China(Grant No.20190402)the Basic Research Foundation of National Commonweal Research Institute of China(Grant No.2017RG004-2).
文摘The effects of putrescine on improving rice growth under aluminum(Al)toxicity conditions have been previously demonstrated,however,the underlying mechanism remains unclear.In this study,treatment with 50 pmol/L Al significantly decreased rice root growth and whole rice dry weight,inhibited Ca2+uptake,decreased ATP syn thesis,and in creased Al,H2O2 and malon dialdehyde(MDA)con tents,whereas the application of putrescine mitigated these negative effects.Putrescine increased root growth and total dry weight of rice,reduced total Al content,decreased H2O2 and MDA contents by increasing antioxidant enzyme(superoxide dismutase,peroxidase,catalase and glutathione S・transferase)activities,increased Ca2+uptake and energy product!oru Proteomic analyses using data-independent acquisition successfully identified 7934 proteins,and 59 representative proteins exhibiting fold-change values higher than 1.5 were randomly selected.From the results of the proteomic and biochemical analyses,we found that putrescine significantly inhibited ethylene biosynthesis and phosphorus uptake in rice roots,increased pectin methylation,decreased pectin content and apoplastic Al deposit!on in rice roots.Putrescine also alleviated Al toxicity by repairing damaged DNA and increasing the proteins involved in maintaining plasma membra ne integrity and normal cell proliferation.These fin dings improve our understanding of how putrescine affects the rice response to Al toxicity,which will facilitate further studies on environmental protection,crop safety,in novations in rice performance and real-world producti on.
基金supported by the National Research Foundation of Korea(NRF)grant funded by the Korean government(MSIT)(Grant Nos.:NRF-2018R1A5A2024425,NRF-2012M3A9C4048796,and NRF-2021R1I1A4A01057387)funded by the National Institutes of Health Office of Research Infrastructure Programs(Grant No.:P40 OD010440)supported by Plant Genomics and Breeding Institute at Seoul National University.
文摘Lipidomics coverage improvement is essential for functional lipid and pathway construction.A powerful approach to discovering organism lipidome is to combine various data acquisitions,such as full scan mass spectrometry(full MS),data-dependent acquisition(DDA),and data-independent acquisition(DIA).Caenorhabditis elegans(C.elegans)is a useful model for discovering toxic-induced metabolism,highthroughput drug screening,and a variety of human disease pathways.To determine the lipidome of C.elegans and investigate lipid disruption from the molecular level to the system biology level,we used integrative data acquisition.The methyl-tert-butyl ether method was used to extract L4 stage C.elegans after exposure to triclosan(TCS),perfluorooctanoic acid,and nanopolystyrene(nPS).Full MS,DDA,and DIA integrations were performed to comprehensively profile the C.elegans lipidome by Q-Exactive Plus MS.All annotated lipids were then analyzed using lipid ontology and pathway analysis.We annotated up to 940 lipids from 20 lipid classes involved in various functions and pathways.The biological investigations revealed that when C.elegans were exposed to nPS,lipid droplets were disrupted,whereas plasma membrane-functionalized lipids were likely to be changed in the TCS treatment group.The nPS treatment caused a significant disruption in lipid storage.Triacylglycerol,glycerophospholipid,and ether class lipids were those primarily hindered by toxicants.Finally,toxicant exposure frequently involved numerous lipid-related pathways,including the phosphoinositide 3-kinase/protein kinase B pathway.In conclusion,an integrative data acquisition strategy was used to characterize the C.elegans lipidome,providing valuable biological insights into hypothesis generation and validation.
基金the National Natural Science Foundation of China(Nos.82003318,81903185,81972916)the“1+X”Program for Clinical Competency Enhancement-Interdisciplinary Innovation Project of the Second Hospital of Dalian Medical University and the Talent Innovation Support Plan of Dalian,China(No.2021RQ008).
文摘Lung cancer produces a high incidence of malignant tumors.There have been many studies on lung cancer using mass spectrometry(MS)technologies.However,most studies have focused on humoral samples.In this work,26 pairs of tissue samples(tumor vs.para-tumor)from patients with lung cancer were analyzed using liquid chromatography-tandem MS(LC-MS/MS)with data-independent acquisition mode.In total,3152 proteins were quantified from tissue samples with high confidence,including 189 up-regulated and 522 down-regulated proteins(tumor vs.para-tumor).In addition,79 and 690 proteins were identified only in para-cancerous samples and cancerous samples,respectively.The results from bio-informatics tools indicated that altered proteins like PEBP1,HRG and LYZ could be ideal reservoirs for screening the potential biomarkers for lung cancer.It is believed these tissue-specific proteomics results will assist in the studies of lung cancer.
基金supported by the National Natural Science Foundation of China(Grant No.81972492)National Science Fund for Young Scholars(Grant No.21904107)+7 种基金Zhejiang Provincial Natural Science Foundation for Distinguished Young Scholars(Grant No.LR19C050001)Hangzhou Agriculture and Society Advancement Program(Grant No.20190101A04)Westlake Startup Grantresearch funds from the National Cancer Centre Singapore and Singapore General Hospital,Singaporethe National Key R&D Program of China(Grant No.2016YFC0901704)Zhejiang Innovation Discipline Project of Laboratory Animal Genetic Engineering(Grant No.201510)the Netherlands Cancer Society(Grant No.NKI 2014-6651)The Netherlands Organization for Scientific Research(NWO)-Middelgroot(Grant No.91116017)
文摘To address the increasing need for detecting and validating protein biomarkers in clinical specimens,mass spectrometry(MS)-based targeted proteomic techniques,including the selected reaction monitoring(SRM),parallel reaction monitoring(PRM),and massively parallel dataindependent acquisition(DIA),have been developed.For optimal performance,they require the fragment ion spectra of targeted peptides as prior knowledge.In this report,we describe a MS pipeline and spectral resource to support targeted proteomics studies for human tissue samples.To build the spectral resource,we integrated common open-source MS computational tools to assemble a freely accessible computational workflow based on Docker.We then applied the workflow to generate DPHL,a comprehensive DIA pan-human library,from 1096 data-dependent acquisition(DDA)MS raw files for 16 types of cancer samples.This extensive spectral resource was then applied to a proteomic study of 17 prostate cancer(PCa)patients.Thereafter,PRM validation was applied to a larger study of 57 PCa patients and the differential expression of three proteins in prostate tumor was validated.As a second application,the DPHL spectral resource was applied to a study consisting of plasma samples from 19 diffuse large B cell lymphoma(DLBCL)patients and 18 healthy control subjects.Differentially expressed proteins between DLBCL patients and healthy control subjects were detected by DIA-MS and confirmed by PRM.These data demonstrate that the DPHL supports DIA and PRM MS pipelines for robust protein biomarker discovery.DPHL is freely accessible at https://www.iprox.org/page/project.html?id=IPX0001400000.
