Research Articles
- Research Article
Probing the sORF-Encoded Peptides of Deinococcus radiodurans in Response to Extreme Stress
In Brief To investigate the SEPs related to extreme stress, the integrated workflow consisting of multiple omics approaches were streamlined. The SEPome of Deinococcus radiodurans containing 109 novel SEPs was described. Among them, SEP068184 exhibited negative regulation of oxidative stress resistance in a comparative phenotypic assay of its mutants. Collectively, the SEPome dataset provides a rich resource for research on the molecular mechanisms of the response to extreme stress in organisms. - Research Article
A Complex Connection Between the Diversity of Human Gastric Mucin O-Glycans, Helicobacter pylori Binding, Helicobacter Infection and Fucosylation
In BriefHelicobacter pylori colonizes the gastric mucus layer of half the world population. Mucin glycans in this mucus layer govern pathogen virulence, growth, and adhesion. The 650 O-glycan structures identified in this study represent a reliable database of structures that are of putative importance for host–pathogen interactions. We further showed that both the enormous gastric O-glycan diversity and avidity for H. pylori are governed by fucosylation and infection-associated O-glycans. - Research Article
A New Monoclonal Antibody Enables BAR Analysis of Subcellular Importin β1 Interactomes
In Brief Proximity biotinylation with a new monoclonal antibody reveals subcellular importin β1 interactomes. - Research Article
The Mechano-Ubiquitinome of Articular Cartilage: Differential Ubiquitination and Activation of a Group of ER-Associated DUBs and ER Stress Regulators
In Brief Using ubiquitomics, we identified a unique ubiquitin signature of articular cartilage upon mechanical injury. A set of deubiquitinases (DUBs) and ER stress regulators including YOD1, ATXN3, RAD23B, VCP/p97, and Ubiquilin 1 were differentially ubiquitinated and their ubiquitination status was linked to enhanced DUB activity and upregulated ER stress response following injury. We observed a rapid increase in global DUB activity in injured and in human osteoarthritic cartilage. Data implicates DUBs as potential targets to modulate injury-induced tissue damage in osteoarthritis. - Research Article
Relative Quantification of Proteins in Formalin-Fixed Paraffin-Embedded Breast Cancer Tissue Using Multiplexed Mass Spectrometry Assays
In Brief The identification of biomarkers is an important challenge in oncology. In this study, we developed a liquid chromatography coupled to multiple reaction monitoring mass spectrometry assay that allows measuring the expression of target proteins in formalin-fixed paraffin-embedded tumor samples, the standard method for biopsy storage in clinical pathology. The method provided quantitative information on 185 proteins from three groups of breast tumors: triple-negative, HER2-overexpressing, and luminal A. In addition to markers such as HER2 or hormone receptors, we identified several proteins which are expressed differentially in triple-negative breast cancer samples. - Research Article
Quantitative Phosphoproteomics Analysis Uncovers PAK2- and CDK1-Mediated Malignant Signaling Pathways in Clear Cell Renal Cell Carcinoma
In Brief Senturk et al. (2022) identified >16,000 phosphopeptides in clear cell Renal Cell Carcinoma tumors, of which 600 were determined to be differentially regulated between tumor and normal adjacent tissues. Furthermore, several oncogenic pathways were determined to be enriched in the tumors such as RAC1 activation, MAPK and VEGF signaling, EGFR signaling, and cytokine signaling. Moreover, the kinase PAK2 was identified as one of the key drivers of tumor migration and invasion, having prognostic impact on the survival of ccRCC patients. - Research Article
Mass Spectrometry and Machine Learning Reveal Determinants of Client Recognition by Antiamyloid Chaperones
In Brief A wide range of proteins can prevent amyloid formation in vitro and in vivo, but the exact nature of these interactions is unclear. By combining machine learning–based structure prediction of protein complexes with mass spectrometry, we can discern the molecular determinants of specific and nonspecific chaperone–client interactions. - Research Article
Improving Statistical Certainty of Glycosylation Similarity between Influenza A Virus Variants Using Data-Independent Acquisition Mass Spectrometry
In Brief It is necessary to sample glycoprotein glycoforms deeply for biological studies. DDA is limited by missing values that result from limited scan speed and stochastic precursor ion selection. DIA samples all glycopeptide glycoforms, but duty cycle is reduced compared with DDA. DIA outperforms DDA for confident assignment of glycopeptides provided target glycoproteins are relatively pure. - Research Article
Ionizing Radiation Drives Key Regulators of Antigen Presentation and a Global Expansion of the Immunopeptidome
In-Brief It has been known that cancer treatment using radiation therapy activates the immune system. Here, we show that radiation increases a signaling component of the immune system called the major histocompatibility complex on tumor cells. This increase makes the tumor more recognizable to T cells whose roles are to specifically find and kill defective or mutated cells. These findings could give physicians a reason to combine radiation therapy with immunotherapies to use the immune effect of radiation for therapeutic benefits. - Research Article
The in vivo Interaction Landscape of Histones H3.1 and H3.3
In Brief Proximity-dependent (BioID) interactome analysis of the histone H3 variants H3.1 and H3.3 identified 608 interaction partners and expanded the characterized H3 interaction landscape. Among the most significant findings were that the chaperone chromatin assembly factor 1, previously thought to be H3.1-specific, interacts with H3.3 throughout the cell cycle. Preferential binding was identified between H3.1 and components of the mitotic machinery and between H3.3 and transcription factors, notably MYC interactors. - Research Article
Comparative Proteomic Analysis Identifies Key Metabolic Regulators of Gemcitabine Resistance in Pancreatic Cancer
In Brief For pancreatic cancer (PDAC) patients, gemcitabine resistance (GemR) represents a major clinical problem. Global quantitative differential proteomic analysis of highly GemR PDAC cells developed from relatively Gem-sensitive PDAC cell lines identified multiple networks within the thousands of quantified proteins that support adaptation to Gem-induced stress, and reveal the complexity of GemR. Drug metabolism pathways were key contributors to GemR, and expression changes in Gem-metabolizing enzymes suggest both their influence upon Gem responsiveness, and drug-targetable vulnerabilities to improve clinical Gem efficacy. - Research Article
High-Throughput Human Complement C3 N-Glycoprofiling Identifies Markers of Early Onset Type 1 Diabetes Mellitus in Children
In Brief Changes in glycosylation are known to modulate inflammatory responses, so glycans are more and more being considered as biomarkers of various diseases. Little is known about complement component C3 N-glycome as proper methods for its analysis are lacking, so we developed a novel high-throughput LC-MS method for C3 plasma enrichment and subsequent glycome analysis. We demonstrated that C3 N-glycan profile is stable in a healthy individual, and results showed that C3 N-glycosylation profile is significantly changed in type 1 diabetes. - Research Article
Mass Spectrometry Identification of Biomarkers in Extracellular Vesicles From Plasmodium vivax Liver Hypnozoite Infections
In Brief Human malaria caused by Plasmodium vivax represents a major barrier to achieve the goal of malaria elimination as this species evolved a dormant liver form called a hypnozoite that can remain latent after the initial infection. Current diagnostic tools are unable to detect asymptomatic patients harboring hypnozoites. We have used a MS proteomics approach to discover biomarkers of latent liver infections associated with circulating extracellular vesicles in an in vivo model of P. vivax liver hypnozoite infections. - Research Article
Functional Diversity and Evolution of the Drosophila Sperm Proteome
In Brief Fertilization is the sine qua non of animal and plant reproduction, a process conserved across the tree of life. During fertilization, a specific subset of male-derived proteins, many of unknown function, enter and are present in the developing egg and zygote. We therefore reinterrogated previous published sperm proteomes using improved purification and fractionation methods and identified >3000 proteins in sperm including a novel exchange of paralogous ribosomal proteins suggesting complex patterns of paralog switching and selectivity during spermatogenesis. - Research Article
Temporal and Sex-Linked Protein Expression Dynamics in a Familial Model of Alzheimer’s Disease
In Brief Alzheimer’s disease still lacks effective therapies. In this study, we provide a longitudinal proteomic characterization of 5XFAD mice hippocampus hoping to identify new clinical intervention targets. Our proteomics approach reveals a striking increase in protein expression in 5XFAD mice overtime, which includes 23 proteins here newly associated with Alzheimer’s disease. Globally, our data reveals initial temporal signatures of DNA damage and senescence transitioning into neuroinflammatory signatures concurrent with disease progression. - Research Article
Spatial Proteomics Reveals Differences in the Cellular Architecture of Antibody-Producing CHO and Plasma Cell–Derived Cells
In Brief We used state-of-the-art protein spatial profiling together with MS-based protein expression analysis and subsequent bioinformatics analysis to elucidate the subcellular proteome organization and organelle architecture of two antibody-secreting cell lines, a CHO and a murine plasma derived cell line (MPC-11). Characterization of differentially expressed and localized proteins and their associated biological pathways allowed a systematic comparison of both cell lines and implicates further targets for CHO cell line engineering. - Research Article
Quantitative Acetylomics Reveals Dynamics of Protein Lysine Acetylation in Mouse Livers During Aging and Upon the Treatment of Nicotinamide Mononucleotide
In Brief Li et al. have performed label-free quantification of the acetylomes in mouse livers with age and upon the treatment of nicotinamide mononucleotide (NMN), revealing previously unknown acetylation events. The datasets characterize not just the dynamics of acetylation with age but also specific acetylation sites as new aging biomarkers. The investigation indicates that the life-beneficial effect of NMN could be partially reflected by the changes in age-related protein acetylation. NMN has mild effects on acetylation previously reported as substrates of Sirtuins. - Research Article
Early Prediction of COVID-19 Patient Survival by Targeted Plasma Multi-Omics and Machine Learning
In Brief During times of hospital admission overload, triage may be required to maximize the number of survivors. Mass spectometry–based proteomic and metabolomic analysis of COVID patients’ blood, collected at the time of admission to the ICU, enabled a prediction of survival versus nonsurvival with 92% accuracy. These analyses, which can be performed on widely available mass spectrometers, have the potential to assist physicians with these difficult decisions. - Research Article
Cardiac Myosin Filaments are Maintained by Stochastic Protein Replacement
In Brief To examine the mechanisms of myosin filament replacement in mouse hearts, we labeled proteins with D3-leucine and a GFP-tag in vivo. We used a combination of high-resolution mass spectrometry, analytical modeling, biochemical fractionations, and fluorescence imaging to demonstrate that individual molecules are stochastically replaced from within these large macromolecular filamentous complexes. - Research Article
Quantitative Proteomics Identifies Proteins Enriched in Large and Small Extracellular Vesicles
In Brief The proteome of large and small extracellular vesicles has been determined with quantitative mass spectrometry. Tetraspanins, ADAMs, and ESCRT proteins, as well as SNAREs and Rab proteins associated with endosomes were enriched in small EVs, whereas ribosomal, mitochondrial, and nuclear proteins, as well as proteins involved in cytokinesis, were enriched in large EVs. Several proteins previously suggested to be enriched in either small or large EVs were validated, and several additional novel protein markers were suggested. - Research Article
Phosphoproteomic Analysis of FLCN Inactivation Highlights Differential Kinase Pathways and Regulatory TFEB Phosphoserines
In Brief Comprehensive phosphoproteomics (pY + pSTY) of FLCNPOS and FLCNNEG human renal cells in conjunction with INKA and with posttranslational modification–based signature enrichment analyses identified FLCN phosphorylation dependencies. FLCN-dependent kinase pathways were investigated through drug experiments and validated in a BHD tumor cell line. This pinpointed RTK-MAPK1/3-RPS6K1/3 as a key axis downstream of FLCN loss. In addition, we show that FLCN loss induces ROS and modulates localization of the TFEB transcription factor by dephosphorylation of specific serines. - Research Article
Normics: Proteomic Normalization by Variance and Data-Inherent Correlation Structure
In Brief Normalization of proteomic data is necessary for quantitative comparison and to improve statistical power. Share, extent, and direction of differential expression are usually unknown. Normalizing with unbalanced or high shares of differential expression can distort the data. Normics computes a ranking list for the selection of a likely invariant protein subset for normalization. It increases sensitivity, specificity, and quantitative accuracy compared to standard normalization alone. Its reversed ranking list provides a filter for highly variant proteins for downstream bioinformatic analyses. - Research Article
A Proteomic Approach Identifies Isoform-Specific and Nucleotide-Dependent RAS Interactions
In Brief Miller et al. presents a comprehensive and quantitative interactome of RAS, a protein found to be a driver of many human cancers. This resource identifies interactors of the active form of RAS (nucleotide-dependent) as well as isoform-specific (KRAS, HRAS, and NRAS) interactors of RAS. Several of the proteins identified were confirmed as being important for cancer cell viability or senescence. Exploring these proteins for further studies may lead to new strategies to treat cancers caused by RAS mutations. - Research Article
MS2Rescore: Data-Driven Rescoring Dramatically Boosts Immunopeptide Identification Rates
In Brief The integration of newly trained immunopeptide MS2PIP models, DeepLC, and Percolator into one software package called MS2Rescore allows for a significant boost in immunopeptide identification rate as well as a substantial increase in specificity. MS2Rescore is search engine-agnostic and unbiased toward HLA types. MS2Rescore, therefore, shows great promise to extend the current neo- and xeno-epitope landscape in existing and future immunopeptidomics experiments. - Research Article
Single-Cell Quantitative Proteomic Analysis of Human Oocyte Maturation Revealed High Heterogeneity in In Vitro–Matured Oocytes
In Brief Here, we performed single-cell quantitative proteomic analysis of human germinal vesicle (GV), in vivo (IVO), and in vitro matured (IVM) oocytes and found low correlation between protein and mRNA levels. IVM oocytes showed higher heterogeneity in protein expression, which is related to the levels of estradiol per mature follicle on trigger day. This study provides a rich resource to characterize the mechanisms of oocyte maturation and to evaluate the quality heterogeneity of IVM oocytes at protein level.
