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Journal of Agricultural and Food... Aug 2022Barley is one of the key cereal grains for malting and brewing industries. However, climate variability and unprecedented weather events can impact barley yield and...
Barley is one of the key cereal grains for malting and brewing industries. However, climate variability and unprecedented weather events can impact barley yield and end-product quality. The genetic background and environmental conditions are key factors in defining the barley proteome content and malting characteristics. Here, we measure the barley proteome and malting characteristics of three barley lines grown in Western Australia, differing in genetic background and growing location, by applying liquid chromatography-mass spectrometry (LC-MS). Using data-dependent acquisition LC-MS, 1571 proteins were detected with high confidence. Quantitative data acquired using sequential window acquisition of all theoretical (SWATH) MS on barley samples resulted in quantitation of 920 proteins. Multivariate analyses revealed that the barley lines' genetics and their growing locations are strongly correlated between proteins and desired traits such as the malt yield. Linking meteorological data with proteomic measurements revealed how high-temperature stress in northern regions affects seed temperature tolerance during malting, resulting in a higher malt yield. Our results show the impact of environmental conditions on the barley proteome and malt characteristics; these findings have the potential to expedite breeding programs and malt quality prediction.
Topics: Hordeum; Phenotype; Plant Breeding; Proteome; Proteomics
PubMed: 35981222
DOI: 10.1021/acs.jafc.2c03816 -
Molecular & Cellular Proteomics : MCP Sep 2023Myeloid-derived suppressor cells (MDSC) are a heterogeneous cell population of incompletely differentiated immune cells. They are known to suppress T cell activity and...
Myeloid-derived suppressor cells (MDSC) are a heterogeneous cell population of incompletely differentiated immune cells. They are known to suppress T cell activity and are implicated in multiple chronic diseases, which make them an attractive cell population for drug discovery. Here, we characterized the baseline proteomes and phospho-proteomes of mouse MDSC differentiated from a progenitor cell line to a depth of 7000 proteins and phosphorylation sites. We also validated the cellular system for drug discovery by recapitulating and identifying known and novel molecular responses to the well-studied MDSC drugs entinostat and mocetinostat. We established a high-throughput drug screening platform using a MDSC/T cell coculture system and assessed the effects of ∼21,000 small molecule compounds on T cell proliferation and IFN-γ secretion to identify novel MDSC modulator. The most promising candidates were validated in a human MDSC system, and subsequent proteomic experiments showed significant upregulation of several proteins associated with the reduction of reactive oxygen species (ROS). Proteome-wide solvent-induced protein stability assays identified Acyp1 and Cd74 as potential targets, and the ROS-reducing drug phenotype was validated by measuring ROS levels in cells in response to compound, suggesting a potential mode of action. We anticipate that the data and chemical tools developed in this study will be valuable for further research on MDSC and related drug discovery.
Topics: Mice; Humans; Animals; Myeloid-Derived Suppressor Cells; High-Throughput Screening Assays; Proteome; Proteomics; Reactive Oxygen Species
PubMed: 37586548
DOI: 10.1016/j.mcpro.2023.100632 -
Emerging Topics in Life Sciences May 2021Plants rapidly respond to environmental fluctuations through coordinated, multi-scalar regulation, enabling complex reactions despite their inherently sessile nature. In... (Review)
Review
Plants rapidly respond to environmental fluctuations through coordinated, multi-scalar regulation, enabling complex reactions despite their inherently sessile nature. In particular, protein post-translational signaling and protein-protein interactions combine to manipulate cellular responses and regulate plant homeostasis with precise temporal and spatial control. Understanding these proteomic networks are essential to addressing ongoing global crises, including those of food security, rising global temperatures, and the need for renewable materials and fuels. Technological advances in mass spectrometry-based proteomics are enabling investigations of unprecedented depth, and are increasingly being optimized for and applied to plant systems. This review highlights recent advances in plant proteomics, with an emphasis on spatially and temporally resolved analysis of post-translational modifications and protein interactions. It also details the necessity for generation of a comprehensive plant cell atlas while highlighting recent accomplishments within the field.
Topics: Mass Spectrometry; Plants; Protein Processing, Post-Translational; Proteome; Proteomics
PubMed: 33620075
DOI: 10.1042/ETLS20200270 -
Mass Spectrometry Reviews Sep 2022The lacrimal film has attracted increasing interest in the last decades as a potential source of biomarkers of physiopathological states, due to its accessibility,... (Review)
Review
The lacrimal film has attracted increasing interest in the last decades as a potential source of biomarkers of physiopathological states, due to its accessibility, moderate complexity, and responsiveness to ocular and systemic diseases. High-performance liquid chromatography-mass spectrometry (LC-MS) has led to effective approaches to tear proteomics, despite the intrinsic limitations in sample amounts. This review focuses on the recent progress in strategy and technology, with an emphasis on the potential for personalized medicine. After an introduction on lacrimal-film composition, examples of applications to biomarker discovery are discussed, comparing approaches based on pooled-sample and single-tear analysis. Then, the most critical steps of the experimental pipeline, that is, tear collection, sample fractionation, and LC-MS implementation, are discussed with reference to proteome-coverage optimization. Advantages and challenges of the alternative procedures are highlighted. Despite the still limited number of studies, tear quantitative proteomics, including single-tear investigation, could offer unique contributions to the identification of low-invasiveness, sustained-accessibility biomarkers, and to the development of personalized approaches to therapy and diagnosis.
Topics: Biomarkers; Mass Spectrometry; Proteome; Proteomics; Tears
PubMed: 33759206
DOI: 10.1002/mas.21691 -
Molecules (Basel, Switzerland) Jan 2021Vascular bundles play important roles in transporting nutrients, growth signals, amino acids, and proteins between aerial and underground tissues. In order to understand... (Review)
Review
Vascular bundles play important roles in transporting nutrients, growth signals, amino acids, and proteins between aerial and underground tissues. In order to understand these sophisticated processes, a comprehensive analysis of the roles of the components located in the vascular tissues is required. A great deal of data has been obtained from proteomic analyses of vascular tissues in plants, which mainly aim to identify the proteins moving through the vascular tissues. Here, different aspects of the phloem and xylem proteins are reviewed, including their collection methods, and their main biological roles in growth, and biotic and abiotic stress responses. The study of vascular proteomics shows great potential to contribute to our understanding of the biological mechanisms related to development and defense in plants.
Topics: Blood Proteins; Phloem; Plant Proteins; Plants; Proteome; Proteomics; Stress, Physiological; Xylem
PubMed: 33514014
DOI: 10.3390/molecules26030667 -
Nature Biotechnology Dec 2021MaxDIA is a software platform for analyzing data-independent acquisition (DIA) proteomics data within the MaxQuant software environment. Using spectral libraries, MaxDIA...
MaxDIA is a software platform for analyzing data-independent acquisition (DIA) proteomics data within the MaxQuant software environment. Using spectral libraries, MaxDIA achieves deep proteome coverage with substantially better coefficients of variation in protein quantification than other software. MaxDIA is equipped with accurate false discovery rate (FDR) estimates on both library-to-DIA match and protein levels, including when using whole-proteome predicted spectral libraries. This is the foundation of discovery DIA-hypothesis-free analysis of DIA samples without library and with reliable FDR control. MaxDIA performs three- or four-dimensional feature detection of fragment data, and scoring of matches is augmented by machine learning on the features of an identification. MaxDIA's bootstrap DIA workflow performs multiple rounds of matching with increasing quality of recalibration and stringency of matching to the library. Combining MaxDIA with two new technologies-BoxCar acquisition and trapped ion mobility spectrometry-both lead to deep and accurate proteome quantification.
Topics: Peptide Library; Proteome; Proteomics; Software
PubMed: 34239088
DOI: 10.1038/s41587-021-00968-7 -
Cell Systems Jan 2021Proteomic technologies now enable the rapid quantification of thousands of proteins across genetically diverse samples. Integration of these data with systems-genetics... (Review)
Review
Proteomic technologies now enable the rapid quantification of thousands of proteins across genetically diverse samples. Integration of these data with systems-genetics analyses is a powerful approach to identify new regulators of economically important or disease-relevant phenotypes in various populations. In this review, we summarize the latest proteomic technologies and discuss technical challenges for their use in population studies. We demonstrate how the analysis of correlation structure and loci mapping can be used to identify genetic factors regulating functional protein networks and complex traits. Finally, we provide an extensive summary of the use of proteome-wide systems genetics throughout fungi, plant, and animal kingdoms and discuss the power of this approach to identify candidate regulators and drug targets in large human consortium studies.
Topics: Humans; Multifactorial Inheritance; Proteome; Proteomics; Research Design
PubMed: 33476553
DOI: 10.1016/j.cels.2020.10.005 -
Analytical Chemistry Jan 2023Large-scale proteome analysis requires rapid and high-throughput analytical methods. We recently reported a new paradigm in proteome analysis where direct infusion and...
Large-scale proteome analysis requires rapid and high-throughput analytical methods. We recently reported a new paradigm in proteome analysis where direct infusion and ion mobility are used instead of liquid chromatography (LC) to achieve rapid and high-throughput proteome analysis. Here, we introduce an improved direct infusion shotgun proteome analysis protocol including label-free quantification (DISPA-LFQ) using CsoDIAq software. With CsoDIAq analysis of DISPA data, we can now identify up to ∼2000 proteins from the HeLa and 293T proteomes, and with DISPA-LFQ, we can quantify ∼1000 proteins from no more than 1 μg of sample within minutes. The identified proteins are involved in numerous valuable pathways including central carbon metabolism, nucleic acid replication and transport, protein synthesis, and endocytosis. Together with a high-throughput sample preparation method in a 96-well plate, we further demonstrate the utility of this technology for performing high-throughput drug analysis in human 293T cells. The total time for data collection from a whole 96-well plate is approximately 8 h. We conclude that the DISPA-LFQ strategy presents a valuable tool for fast identification and quantification of proteins in complex mixtures, which will power a high-throughput proteomic era of drug screening, biomarker discovery, and clinical analysis.
Topics: Humans; Proteome; Proteomics; Chromatography, Liquid; Software
PubMed: 36527718
DOI: 10.1021/acs.analchem.2c02249 -
Journal of Proteome Research Dec 2014Mass spectrometry plays a key role in relative quantitative comparisons of proteins in order to understand their functional role in biological systems upon perturbation.... (Review)
Review
Mass spectrometry plays a key role in relative quantitative comparisons of proteins in order to understand their functional role in biological systems upon perturbation. In this review, we review studies that examine different aspects of isobaric labeling-based relative quantification for shotgun proteomic analysis. In particular, we focus on different types of isobaric reagents and their reaction chemistry (e.g., amine-, carbonyl-, and sulfhydryl-reactive). Various factors, such as ratio compression, reporter ion dynamic range, and others, cause an underestimation of changes in relative abundance of proteins across samples, undermining the ability of the isobaric labeling approach to be truly quantitative. These factors that affect quantification and the suggested combinations of experimental design and optimal data acquisition methods to increase the precision and accuracy of the measurements will be discussed. Finally, the extended application of isobaric labeling-based approach in hyperplexing strategy, targeted quantification, and phosphopeptide analysis are also examined.
Topics: Amines; Isotope Labeling; Mass Spectrometry; Phosphopeptides; Protein Carbonylation; Proteome; Proteomics; Reproducibility of Results; Sulfhydryl Compounds
PubMed: 25337643
DOI: 10.1021/pr500880b -
Cells Aug 2022Dissecting the proteome of cell types and states at single-cell resolution, while being highly challenging, has significant implications in basic science and...
Dissecting the proteome of cell types and states at single-cell resolution, while being highly challenging, has significant implications in basic science and biomedicine. Mass spectrometry (MS)-based single-cell proteomics represents an emerging technology for system-wide, unbiased profiling of proteins in single cells. However, significant challenges remain in analyzing an extremely small amount of proteins collected from a single cell, as a proteome-wide amplification of proteins is not currently feasible. Here, we report an integrated spectral library-based single-cell proteomics (SLB-SCP) platform that is ultrasensitive and well suited for a large-scale analysis. To overcome the low MS/MS signal intensity intrinsically associated with a single-cell analysis, this approach takes an alternative approach by extracting a breadth of information that specifically defines the physicochemical characteristics of a peptide from MS1 spectra, including monoisotopic mass, isotopic distribution, and retention time (hydrophobicity), and uses a spectral library for proteomic identification. This conceptually unique MS platform, coupled with the DIRECT sample preparation method, enabled identification of more than 2000 proteins in a single cell to distinguish different proteome landscapes associated with cellular types and heterogeneity. We characterized individual normal and cancerous pancreatic ductal cells (HPDE and PANC-1, respectively) and demonstrated the substantial difference in the proteomes between HPDE and PANC-1 at the single-cell level. A significant upregulation of multiple protein networks in cancer hallmarks was identified in the PANC-1 cells, functionally discriminating the PANC-1 cells from the HPDE cells. This integrated platform can be built on high-resolution MS and widely accepted proteomic software, making it possible for community-wide applications.
Topics: Peptides; Proteome; Proteomics; Software; Tandem Mass Spectrometry
PubMed: 35954294
DOI: 10.3390/cells11152450