Research Articles
- Research Article Collection: Cancer Proteomics
Deciphering the Clinical Significance and Kinase Functions of GSK3α in Colon Cancer by Proteomics and Phosphoproteomics
In Brief GSK3α and GSK3β are distinct isoforms, with GSK3β studied in cancer development while GSK3α was considered redundant. Yet, our research has revealed that GSK3α, rather than GSK3β, is significantly correlated with colon cancer patients' survival. Integrative In vitro and in cell line phosphoproteomics identified 156 phosphosites specifically regulated by GSK3α. Notably, phosphorylation of THRAP3S248 remarkably enhanced colon cancer cell migration. These findings suggest that GSK3α may have unique and important roles in colon cancer development and progression. - Research Article
N-Terminomic Changes in Neurons During Excitotoxicity Reveal Proteolytic Events Associated With Synaptic Dysfunctions and Potential Targets for Neuroprotection
In Brief Using N-terminomics and global proteomics approaches, Ameen et al. mapped changes in protein abundance and N-terminome of cultured primary neurons during excitotoxicity, a crucial neuronal death process in neurological disorders. These proteomic changes document new excitotoxicity-associated molecular events and offer insights into how these events are organized to induce neuronal death. The potential therapeutic relevance of these molecular events is illustrated by the demonstration that in vivo blockade of one of these events could protect against excitotoxic neuronal loss. - Research Article
Translational and Posttranslational Dynamics in a Model Peptidergic System
In Brief In this work, we have explored the proteomes and phosphoproteomes of somatic neuronal compartments in the supraoptic nucleus of the hypothalamus and their axonal projections in the hypophysis under basal and stimulated conditions. Through comparisons with corresponding transcriptomes of the supraoptic nucleus, we bring a novel perspective to transcriptome, proteome, and phosphoproteome dynamics in this uniquely tractable model neuronal system. - Research Article
Integrative Proteomics and N-Glycoproteomics Analyses of Rheumatoid Arthritis Synovium Reveal Immune-Associated Glycopeptides
In Brief Protein glycosylation plays key roles in the pathogenesis of RA but in-depth glycoproteomics analysis of synovial tissues is still lacking. Here, we established the global N-glycoproteomics landscape of synovium from RA and OA patients by using a TMT-based strategy. Our results revealed that the hyper-glycosylated proteins in RA were closely linked to immune responses. We identified some N-glycopeptides correlated with the filtration of certain immune cells. This work identifies immune-associated N-glycoproteomics signatures of RA synovium. - Research Article
Quantitative Proteomic Analysis Reveals apoE4-Dependent Phosphorylation of the Actin-Regulating Protein VASP
In Brief Apolipoprotein (apo) E4 is the major genetic risk factor for Alzheimer’s Disease (AD). Here, we measured the impact of apoE4 on protein phosphorylation and ubiquitylation. ApoE4 expression resulted in a dramatic increase in VASP S235 phosphorylation in a PKA-dependent manner resulting in the disruption of VASP interactions with numerous actin cytoskeletal and microtubular proteins. Reduction of VASP S235 phosphorylation via PKA inhibition resulted in a significant increase in neurite outgrowth in apoE4-expressing cells. - Research Article
MSFragger-Labile: A Flexible Method to Improve Labile PTM Analysis in Proteomics
In Brief Many peptide modifications are labile or fragment during tandem mass spectrometry experiments for proteomics. Database search engines often struggle to identify peptides bearing labile modifications, as the expected masses of fragment ions (with intact modification(s)) do not match the observed spectra. We have developed MSFragger Labile, a flexible set of parameters that incorporate modification fragmentation information into MSFragger search. We show that labile mode can greatly increase the number of modified peptides identified by database search. - Research Article Collection: Cancer Proteomics
Data-Independent Acquisition Phosphoproteomics of Urinary Extracellular Vesicles Enables Renal Cell Carcinoma Grade Differentiation
In Brief Considering the limitations of current approaches, it is necessary to develop a novel diagnostic technique for early intervention of renal cell carcinoma (RCC). Here, we made our efforts to adapt the existing DIA method to analyze urinary EV phosphoproteomics for noninvasive RCC biomarker screening. Combined with our in-house EVtrap method and PolyMAC enrichment, we quantified 2584 unique phosphosites. We observed unique upregulated phosphosites and pathways differentiating healthy control (HC), chronic kidney disease (CKD), low-grade, and high-grade clear cell RCC. - Research Article
Proteomic Analysis of Huntington’s Disease Medium Spiny Neurons Identifies Alterations in Lipid Droplets
In Brief The proteome of the medium spiny neurons of human Huntington’s disease from isogenic HD iPSCs is defined and potential therapeutic targets or biomarkers are defined. - Research Article
Development of a Spectral Library for the Discovery of Altered Genomic Events in Mycobacterium avium Associated With Virulence Using Mass Spectrometry–Based Proteogenomic Analysis
In Brief Detection of coinfecting pathogens in clinical samples is challenging owing to lack of tools and molecular data. Through a combination of in-depth proteomics and integrated genomic analysis, we identified several unique molecular leads in Mycobacterium avium with a potential to distinguish it from other coinfecting pathogens in a unique mass spectrometry–based data-independent acquisition assay. As and when more molecular data emanate from other coinfecting pathogens are available, this assay can be expanded for the detection of multiple pathogens. - Research Article Collection: Cancer Proteomics
Phenotypic Heterogeneity Analysis of APC-Mutant Colon Cancer by Proteomics and Phosphoproteomics Identifies RAI14 as a Key Prognostic Determinant in East Asians and Westerners
In Brief The prognostic phenotype is heterogeneous in colorectal cancer (CRC) patients with APC mutations, while the diagnostic biomarkers remain largely unknown. Here, we described the phenotypic heterogeneity of APC-mutant tumors and identified RAI14 as a key prognostic determinant for APC-mutant colon cancer patients. The prognostic utility of RAI14 in APC-mutant colon cancer will provide early warning and increase the chance of successful treatment. - Research Article
PI3Kα Translocation Mediates Nuclear PtdIns(3,4,5)P3 Effector Signaling in Colorectal Cancer
In Brief The canonical view of PI3Kα signaling describes PtdIns(3,4,5)P3 generation and activation of downstream effectors at the plasma membrane. Here, we show that colorectal cancer cell lines exhibit a diverse plasma membrane–nuclear distribution of PI3Kα, controlling corresponding subcellular PtdIns(3,4,5)P3 pools, and characterize the nuclear PtdIns(3,4,5)P3 interactome. Our findings support a model in which nuclear translocation of PI3Kα defines a mechanism for spatial organization of PtdIns(3,4,5)P3 effector signaling, associated with colorectal cancer subtypes, and modulating signaling responses. - Research Article
Characterization of p38α Signaling Networks in Cancer Cells Using Quantitative Proteomics and Phosphoproteomics
In Brief We have used proteomic and phosphoproteomic analyses to study the rewiring of signaling pathways in breast cancer cells upon interfering with p38α function. The identification of a set of differentially expressed proteins and phosphorylation changes allowed us to propose a network of protein kinases and cellular processes regulated by p38α. We also show that p38α can control cancer cell adhesion through the modulation of the protein ArgBP2. - Research Article
Integrated MicroRNA and Secretome Analysis of Human Endometrial Organoids Reveal the miR-3194-5p/Aquaporin/S100A9 Module in Regulating Trophoblast Functions
In Brief In this study, we adopted the human endometrial organoid model to simulate the endometrium gland responses under the hormonal conditions in the menstrual cycle and early gestation. This study filled the research gap of the miRNA expression and secretome profiles of endometrial organoids in the estrous cycle and early pregnancy stages. Secretome of endometrial organoids treated with hormones mimicking early pregnancy enhances trophoblast invasion and migration, which is mediated by the miR-3194-5p/AQP-1&AQP-9/S100A9 regulatory axis. - Research Article Collection: Neurobiology Proteomics
Cold Exposure–induced Alterations in the Brain Peptidome and Gut Microbiome Are Linked to Energy Homeostasis in Mice
In Brief To interrogate the interplay between brain peptides and gut microbes in response to cold, we performed a region-resolvable quantitative profiling of the brain peptidome using cold-exposed mouse models. ProSAAS-derived peptides exhibited positive correlation with Lactobacillus. The hypothalamus–pituitary axis exhibited a sensitive response. Intervention with cold-adapted microbiota in mice decreased the abundance of hypothalamic neurokinin B and subsequently contributed to energy consumption change. This study provides a data resource for understanding the regulatory mechanism of energy homeostasis upon cold exposure. - Research Article Collection: Virology Proteomics
Cerebrospinal Fluid Protein Markers Indicate Neuro-Damage in SARS-CoV-2-Infected Nonhuman Primates
In Brief Proteomic quantitation of CSF samples from control and SARS-CoV-2-infected nonhuman primates to identify differentially expressed proteins to understand regulatory pathways associated with neuropathophysiology after SARS-CoV-2 infection. - Research Article
Serum Proteomics Identifies Biomarkers Associated With the Pathogenesis of Idiopathic Pulmonary Fibrosis
In Brief Serum proteomics distinguished IPF patients into three subgroups in signal pathways and overall survival. Aging-associated signatures provided clear and direct evidence that aging is a critical risk factor for IPF rather than to a single biomarker. LDHA and CCT6A expression, which were associated with glucose metabolic reprogramming, were correlated with high serum lactic acid content in IPF patients. Cross-model analysis and machine learning showed that the combinatorial biomarker is applicable and validated from another cohort and ELISA assay. - Research Article
Chemical Genetic Identification of PKC Epsilon Substrates in Mouse Brain
In Brief PKC epsilon (PKCε) plays important roles in behavioral responses to alcohol and in anxiety-like behavior, making it a potential drug target for reducing alcohol consumption and anxiety. Here, a chemical genetic screen and mass spectrometry identified 39 direct substrates of PKCε in mouse brain. They were prioritized using several public databases (LINCS-L1000, STRING, GeneFriends, GeneMANIA) to predict interactions between them and PKCε. Many were novel and broadly fell into three functional categories: cytoskeletal regulation, morphogenesis, and synaptic function. - Research Article
cKMT1 is a New Lysine Methyltransferase That Methylates the Ferredoxin-NADP(+) Oxidoreductase and Regulates Energy Transfer in Cyanobacteria
In Brief We biochemically identified a new lysine methyltransferase (cKMT1) in a model cyanobacterium Synechocystis. The loss of cKMT1 affects photosynthetic electron transfer in Synechocystis. Quantitative analysis of the lysine methylation identified 58 endogenous substrates of cKMT1. We demonstrated that cKMT1 can methylate ferredoxin-NADP(+) oxidoreductase (FNR) to regulate its enzymatic activity and affect photosynthetic energy transfer. This study revealed a methylation-mediated molecular mechanism catalyzed by cKMT1 for the regulation of photosynthesis in cyanobacteria. - Research Article
Serum Exosomes From Epithelial Ovarian Cancer Patients Contain LRP1, Which Promotes the Migration of Epithelial Ovarian Cancer Cell
In Brief Ovarian cancer is the most lethal malignant gynecological tumor, and the clinical diagnosis and treatment of OC have always been difficult. Cancer cell–derived exosomes play a key role in mediating cancer progression. We performed proteomics analysis of serum exosomes to discover biomarkers for epithelial ovarian cancer. - Research Article Collection: Single-cell Proteomics
Challenges and Opportunities for Single-cell Computational Proteomics
In Brief Single-cell proteomics is growing rapidly, but most research has focused on improving instrumentation and sample preparation. Current algorithms being employed on single-cell data were designed for bulk data and have underlying assumptions that may not hold true for single-cell data. Here, we present a review of algorithms for identifying and quantifying peptides and proteins, including how each type of algorithm works, assumptions it relies on, and possible optimizations for single-cell data. - Research Article
Large-Scale Immunopeptidome Analysis Reveals Recurrent Posttranslational Splicing of Cancer- and Immune-Associated Genes
In Brief Thus far, no consensus has been reached on the extent to which post-translational spliced peptides occur, stirring significant debate. Here, we designed a pipeline for their identification. Key strategies for the filtration of noncanonical peptides are proposed following comparative filtration to WT peptides. We find that only low percentages of the spliced peptides are identified for HLA class-I and II. Spliced peptides are further validated based on synthetic peptides and immune-reactivity assays. - Research Article
Comprehensive Proteomics Analysis Identifies CD38-Mediated NAD+ Decline Orchestrating Renal Fibrosis in Pediatric Patients With Obstructive Nephropathy
In Brief Obstructive nephropathy is a leading cause of kidney injury in infants and children. In this work, we performed comparative proteomics of control and obstructed kidneys from human and experimental obstructive nephropathy and uncovered the aberrant NAD+ metabolism, which was partially induced by CD38 upregulation. Deletion or inhibition of CD38 reduced obstruction-associated renal fibrosis and inflammation. These findings emphasized the therapeutic potential of CD38 and NAD+ metabolism in obstructive nephropathy. - Research Article
The Proteome of Large or Small Extracellular Vesicles in Pig Seminal Plasma Differs, Defining Sources and Biological Functions
In Brief Pig seminal plasma contains a heterogeneous population of extracellular vesicles (sEVs), involved in several reproductive processes. Two sEV subsets (small or large EVs) were isolated by size exclusion chromatography (SEC) followed by a large-scale high-throughput proteomic analysis. A total of 1,034 proteins were identified by LC-MS/MS as encoded into Sus scrofa taxonomy, and 737 of them were quantified by SWATH revealing quantitative proteomic differences that suggest different biogenesis and biological functions for the sEVs. - Research Article Collection: Immunopeptidomics
Benchmarking Bioinformatics Pipelines in Data-Independent Acquisition Mass Spectrometry for Immunopeptidomics
In Brief We benchmarked four commonly used “peptide-centric” software tools—Skyline, Spectronaut, DIA-NN, and PEAKS—for spectral library-based data-independent acquisition analysis of immunopeptidomics data. - Research Article
Precision Neoantigen Discovery Using Large-Scale Immunopeptidomes and Composite Modeling of MHC Peptide Presentation
In Brief Accurately identifying neoantigens is critical for many clinical applications. We generated immunopeptidomics data from 25 stably transfected monoallelic cell lines. Then, we systematically reprocessed a large corpus of public data to improve MHC binding pocket diversity and to empirically learn the rules of antigen presentation. In applying these datasets, we trained SHERPA, an MHC binding and presentation prediction algorithm. SHERPA improves performance compared with existing tools by 1.44-fold in held-out monoallelic data and 1.11-fold for known immunogenic epitopes.
