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Thrombosis and Haemostasis Jul 2022Proteomics, the simultaneous study of all proteins in a given cell, tissue or organism, is an innovative approach used to identify novel markers for diagnosis, prognosis... (Review)
Review
Proteomics, the simultaneous study of all proteins in a given cell, tissue or organism, is an innovative approach used to identify novel markers for diagnosis, prognosis and the pathophysiological mechanisms associated with diseases. Proteomic methodologies have been used in a variety of contexts such as investigating changes in protein abundance that may occur with disease presence, the response to therapeutic treatments as well as the impacts of age on the plasma proteome.Over the last decade, significant technological advancements in proteomic techniques have resulted in an increase in the use of proteomics in thrombosis and hemostasis research, particularly in order to identify relevant and novel clinical markers associated with bleeding and thrombosis. This mini-review explores the use of proteomics in the setting of thrombosis and hemostasis from 2010-2020, across five main domains (platelets, blood clot composition, stroke, venous thromboembolism, and therapeutics), as well as provides insights into key considerations for conducting proteomic studies.
Topics: Biomarkers; Blood Platelets; Hemostasis; Humans; Proteome; Proteomics; Thrombosis
PubMed: 34753192
DOI: 10.1055/a-1690-8897 -
Molecular Cancer Feb 2022Alterations in DNAs could not reveal what happened in proteins. The accumulated alterations of DNAs would change the manifestation of proteins. Therefore, as is the case... (Review)
Review
Alterations in DNAs could not reveal what happened in proteins. The accumulated alterations of DNAs would change the manifestation of proteins. Therefore, as is the case in cancer liquid biopsies, deep proteome profiling will likely provide invaluable and clinically relevant information in real-time throughout all stages of cancer progression. However, due to the great complexity of proteomes in liquid biopsy samples and the limitations of proteomic technologies compared to high-plex sequencing technologies, proteomic discoveries have yet lagged behind their counterpart, genomic technologies. Therefore, novel protein technologies are in urgent demand to fulfill the goals set out for biomarker discovery in cancer liquid biopsies.Notably, conventional and innovative technologies are being rapidly developed for proteomic analysis in cancer liquid biopsies. These advances have greatly facilitated early detection, diagnosis, prognosis, and monitoring of cancer evolution, adapted or adopted in response to therapeutic interventions. In this paper, we review the high-plex proteomics technologies that are capable of measuring at least hundreds of proteins simultaneously from liquid biopsy samples, ranging from traditional technologies based on mass spectrometry (MS) and antibody/antigen arrays to innovative technologies based on aptamer, proximity extension assay (PEA), and reverse phase protein arrays (RPPA).
Topics: Early Detection of Cancer; Humans; Liquid Biopsy; Neoplasms; Proteome; Proteomics
PubMed: 35168611
DOI: 10.1186/s12943-022-01526-8 -
Cancer Cell Aug 2022The proteome provides unique insights into disease biology beyond the genome and transcriptome. A lack of large proteomic datasets has restricted the identification of...
The proteome provides unique insights into disease biology beyond the genome and transcriptome. A lack of large proteomic datasets has restricted the identification of new cancer biomarkers. Here, proteomes of 949 cancer cell lines across 28 tissue types are analyzed by mass spectrometry. Deploying a workflow to quantify 8,498 proteins, these data capture evidence of cell-type and post-transcriptional modifications. Integrating multi-omics, drug response, and CRISPR-Cas9 gene essentiality screens with a deep learning-based pipeline reveals thousands of protein biomarkers of cancer vulnerabilities that are not significant at the transcript level. The power of the proteome to predict drug response is very similar to that of the transcriptome. Further, random downsampling to only 1,500 proteins has limited impact on predictive power, consistent with protein networks being highly connected and co-regulated. This pan-cancer proteomic map (ProCan-DepMapSanger) is a comprehensive resource available at https://cellmodelpassports.sanger.ac.uk.
Topics: Biomarkers, Tumor; Cell Line; Humans; Neoplasms; Proteome; Proteomics
PubMed: 35839778
DOI: 10.1016/j.ccell.2022.06.010 -
International Journal of Molecular... Mar 2023Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics is a powerful technique for profiling proteomes of cells, tissues, and body fluids. Typical... (Review)
Review
Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics is a powerful technique for profiling proteomes of cells, tissues, and body fluids. Typical bottom-up proteomic workflows consist of the following three major steps: sample preparation, LC-MS/MS analysis, and data analysis. LC-MS/MS and data analysis techniques have been intensively developed, whereas sample preparation, a laborious process, remains a difficult task and the main challenge in different applications. Sample preparation is a crucial stage that affects the overall efficiency of a proteomic study; however, it is prone to errors and has low reproducibility and throughput. In-solution digestion and filter-aided sample preparation are the typical and widely used methods. In the past decade, novel methods to improve and facilitate the entire sample preparation process or integrate sample preparation and fractionation have been reported to reduce time, increase throughput, and improve reproducibility. In this review, we have outlined the current methods used for sample preparation in proteomics, including on-membrane digestion, bead-based digestion, immobilized enzymatic digestion, and suspension trapping. Additionally, we have summarized and discussed current devices and methods for integrating different steps of sample preparation and peptide fractionation.
Topics: Chromatography, Liquid; Tandem Mass Spectrometry; Proteomics; Reproducibility of Results; Peptides; Proteome
PubMed: 36982423
DOI: 10.3390/ijms24065350 -
Cell Reports. Medicine Jun 2022Parkinson's disease (PD) is a growing burden worldwide, and there is no reliable biomarker used in clinical routines to date. Cerebrospinal fluid (CSF) is routinely...
Parkinson's disease (PD) is a growing burden worldwide, and there is no reliable biomarker used in clinical routines to date. Cerebrospinal fluid (CSF) is routinely collected in patients with neurological symptoms and should closely reflect alterations in PD patients' brains. Here, we describe a scalable and sensitive mass spectrometry (MS)-based proteomics workflow for CSF proteome profiling. From two independent cohorts with over 200 individuals, our workflow reproducibly quantifies over 1,700 proteins from minimal CSF amounts. Machine learning determines OMD, CD44, VGF, PRL, and MAN2B1 to be altered in PD patients or to significantly correlate with clinical scores. We also uncover signatures of enhanced neuroinflammation in LRRK2 G2019S carriers, as indicated by increased levels of CTSS, PLD4, and HLA proteins. A comparison with our previously acquired urinary proteomes reveals a large overlap in PD-associated changes, including lysosomal proteins, opening up new avenues to improve our understanding of PD pathogenesis.
Topics: Biomarkers; Heterozygote; Humans; Parkinson Disease; Proteome; Proteomics
PubMed: 35732154
DOI: 10.1016/j.xcrm.2022.100661 -
Nature Mar 2003The sequencing of the human genome and that of numerous pathogens has opened the door for proteomics by providing a sequence-based framework for mining proteomes. As a... (Review)
Review
The sequencing of the human genome and that of numerous pathogens has opened the door for proteomics by providing a sequence-based framework for mining proteomes. As a result, there is intense interest in applying proteomics to foster a better understanding of disease processes, develop new biomarkers for diagnosis and early detection of disease, and accelerate drug development. This interest creates numerous opportunities as well as challenges to meet the needs for high sensitivity and high throughput required for disease-related investigations.
Topics: Disease; Drug Design; Electrophoresis, Gel, Two-Dimensional; Gene Expression Profiling; Humans; Neoplasms; Oligonucleotide Array Sequence Analysis; Protein Array Analysis; Proteome; Proteomics
PubMed: 12634796
DOI: 10.1038/nature01514 -
Nature Biotechnology Dec 2017Although advances in protein labeling methods have made it possible to measure the proteome of mixed cell populations, it has not been possible to isolate...
Although advances in protein labeling methods have made it possible to measure the proteome of mixed cell populations, it has not been possible to isolate cell-type-specific proteomes in vivo. This is because the existing methods for metabolic protein labeling in vivo access all cell types. We report the development of a transgenic mouse line where Cre-recombinase-induced expression of a mutant methionyl-tRNA synthetase (L274G) enables the cell-type-specific labeling of nascent proteins with a non-canonical amino-acid and click chemistry. Using immunoblotting, imaging and mass spectrometry, we use our transgenic mouse to label and analyze proteins in excitatory principal neurons and Purkinje neurons in vitro (brain slices) and in vivo. We discover more than 200 proteins that are differentially regulated in hippocampal excitatory neurons by exposing mice to an environment with enriched sensory cues. Our approach can be used to isolate, analyze and quantitate cell-type-specific proteomes and their dynamics in healthy and diseased tissues.
Topics: Amino Acids; Animals; Click Chemistry; Female; Gene Expression Regulation; Integrases; Male; Methionine-tRNA Ligase; Mice; Mice, Transgenic; Neurons; Proteome; Proteomics
PubMed: 29106408
DOI: 10.1038/nbt.4016 -
Molecular Systems Biology Mar 2022Single-cell technologies are revolutionizing biology but are today mainly limited to imaging and deep sequencing. However, proteins are the main drivers of cellular...
Single-cell technologies are revolutionizing biology but are today mainly limited to imaging and deep sequencing. However, proteins are the main drivers of cellular function and in-depth characterization of individual cells by mass spectrometry (MS)-based proteomics would thus be highly valuable and complementary. Here, we develop a robust workflow combining miniaturized sample preparation, very low flow-rate chromatography, and a novel trapped ion mobility mass spectrometer, resulting in a more than 10-fold improved sensitivity. We precisely and robustly quantify proteomes and their changes in single, FACS-isolated cells. Arresting cells at defined stages of the cell cycle by drug treatment retrieves expected key regulators. Furthermore, it highlights potential novel ones and allows cell phase prediction. Comparing the variability in more than 430 single-cell proteomes to transcriptome data revealed a stable-core proteome despite perturbation, while the transcriptome appears stochastic. Our technology can readily be applied to ultra-high sensitivity analyses of tissue material, posttranslational modifications, and small molecule studies from small cell counts to gain unprecedented insights into cellular heterogeneity in health and disease.
Topics: Mass Spectrometry; Protein Processing, Post-Translational; Proteome; Proteomics; Workflow
PubMed: 35226415
DOI: 10.15252/msb.202110798 -
Journal of Proteomics Oct 2018The enormous diversity of proteoforms produces tremendous complexity within cellular proteomes, facilitates intricate networks of molecular interactions, and constitutes... (Review)
Review
The enormous diversity of proteoforms produces tremendous complexity within cellular proteomes, facilitates intricate networks of molecular interactions, and constitutes a formidable analytical challenge for biomedical researchers. Currently, quantitative whole-proteome profiling often relies on non-targeted liquid chromatography-mass spectrometry (LC-MS), which samples proteoforms broadly, but can suffer from lower accuracy, sensitivity, and reproducibility compared with targeted LC-MS. Recent advances in bottom-up proteomics using targeted LC-MS have enabled previously unachievable identification and quantification of target proteins and posttranslational modifications within complex samples. Consequently, targeted LC-MS is rapidly advancing biomedical research, especially systems biology research in diverse areas that include proteogenomics, interactomics, kinomics, and biological pathway modeling. With the recent development of targeted LC-MS assays for nearly the entire human proteome, targeted LC-MS is positioned to enable quantitative proteomic profiling of unprecedented quality and accessibility to support fundamental and clinical research. Here we review recent applications of bottom-up proteomics using targeted LC-MS for systems biology research. SIGNIFICANCE: Advances in targeted proteomics are rapidly advancing systems biology research. Recent applications include systems-level investigations focused on posttranslational modifications (such as phosphoproteomics), protein conformation, protein-protein interaction, kinomics, proteogenomics, and metabolic and signaling pathways. Notably, absolute quantification of metabolic and signaling pathway proteins has enabled accurate pathway modeling and engineering. Integration of targeted proteomics with other technologies, such as RNA-seq, has facilitated diverse research such as the identification of hundreds of "missing" human proteins (genes and transcripts that appear to encode proteins but direct experimental evidence was lacking).
Topics: Animals; Biomedical Research; Gene Expression Profiling; Humans; Mass Spectrometry; Protein Processing, Post-Translational; Proteome; Proteomics; Signal Transduction; Systems Biology
PubMed: 29452276
DOI: 10.1016/j.jprot.2018.02.008 -
Cell Nov 2023Neurons build synaptic contacts using different protein combinations that define the specificity, function, and plasticity potential of synapses; however, the diversity...
Neurons build synaptic contacts using different protein combinations that define the specificity, function, and plasticity potential of synapses; however, the diversity of synaptic proteomes remains largely unexplored. We prepared synaptosomes from 7 different transgenic mouse lines with fluorescently labeled presynaptic terminals. Combining microdissection of 5 different brain regions with fluorescent-activated synaptosome sorting (FASS), we isolated and analyzed the proteomes of 18 different synapse types. We discovered ∼1,800 unique synapse-type-enriched proteins and allocated thousands of proteins to different types of synapses (https://syndive.org/). We identify shared synaptic protein modules and highlight the proteomic hotspots for synapse specialization. We reveal unique and common features of the striatal dopaminergic proteome and discover the proteome signatures that relate to the functional properties of different interneuron classes. This study provides a molecular systems-biology analysis of synapses and a framework to integrate proteomic information for synapse subtypes of interest with cellular or circuit-level experiments.
Topics: Animals; Mice; Brain; Mice, Transgenic; Proteome; Proteomics; Synapses; Synaptosomes
PubMed: 37918396
DOI: 10.1016/j.cell.2023.09.028