Acquiring and Analyzing Data Independent Acquisition Proteomics Experiments without Spectrum Libraries

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Highlights

• •Rapid DIA-only library building with gas-phase fractionation.
• •Recommended DIA acquisition strategies with staggered windows and forbidden zones.
• •Optimized DIA instrument settings for several Thermo Orbitrap instruments.
• •Data analysis tutorial using open source DIA software.

     

MaxQuant Software for Ion Mobility Enhanced Shotgun Proteomics

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Highlights

• •Highly parallelizable 4D feature detection in ion mobility enhanced shotgun proteomics.
• •Multidimensional non-linear mass, retention time and ion mobility recalibration.
• •Collision cross section aware matching between runs.
• •Label-free quantification of ion mobility MS data.

     

A Cross-linking Mass Spectrometry Approach Defines Protein Interactions in Yeast Mitochondria

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Highlights

• •XL-MS reveals new PPIs in yeast mitochondria under glycerol and glucose condition.
• •Significant but limited results from quantitative XL-MS experiments.
• •Ndi1 participates in a CIII₂CIV₂ respiratory supercomplex.
• •Min8 promotes assembly of Cox12 into an intermediate complex IV.

     

Selection of Features with Consistent Profiles Improves Relative Protein Quantification in Mass Spectrometry Experiments

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Highlights

• •Automated statistical approach for detecting uninformative features and outliers.
• •Improved performance on relative protein quantification.
• •An option in the open-source R-based software MSstats.

     

Chemical Genetics of AGC-kinases Reveals Shared Targets of Ypk1, Protein Kinase A and Sch9

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Highlights

• •Acute inhibition of growth-regulatory AGC-kinases Sch9, PKA and Ypk1.
• •Phospho-proteomic profiling of 6373 phsopho-sites.
• •Ypk1 regulates phospho-sites in RRxS/T-context and functionally overlaps with PKA.
• •Ypk1 and PKA activate trehalase activity of Nth1.

     

Robust Summarization and Inference in Proteome-wide Label-free Quantification

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Highlights

• •In depth performance assessment of leading tools for differential protein abundance.
• •Novel fast modular framework MSqRobSum for robust protein summarization and inference.
• •MsqRobSum outperforms leading protein summarization-based tools.
• •MSqRobSum is on par with top-performing peptide based tool MSqRob.

     

DEqMS: A Method for Accurate Variance Estimation in Differential Protein Expression Analysis

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Highlights

• •DEqMS is a method for statistical analysis of quantitative MS-data.
• •Variance estimates based on the actual MS-data structure.
• •Improved statistical power and accuracy in protein differential analysis.
• •DEqMS is available as a user-friendly R package in Bioconductor.

     

An Improved Boosting to Amplify Signal with Isobaric Labeling (iBASIL) Strategy for Precise Quantitative Single-cell Proteomics

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Highlights

• •Higher AGC significantly improves quantitation quality in single-cell analysis.
• •The boosting-to-sample ratio should be carefully evaluated and optimized.
• •iBASIL allows for precise quantitation of 1,500 proteins from 104 AML single cells.
• •iBASIL recapitulates major biological differences in different AML single cells.

     

Tandem Mass Tag Approach Utilizing Pervanadate BOOST Channels Delivers Deeper Quantitative Characterization of the Tyrosine Phosphoproteome

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Highlights

• •Detection of low-abundance phosphotyrosine-containing peptides is challenging.
• •Multiplexed TMT allows inclusion of modification(pTyr)-saturated boost channels.
• •Boost channels facilitate selection of pTyr precursor ions for fragmentation.
• •Quantitation depth is increased while maintaining accuracy and precision.

     

Fast Quantitative Analysis of timsTOF PASEF Data with MSFragger and IonQuant

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Highlights

• •MSFragger now supports raw timsTOF PASEF data.
• •IonQuant performs fast and accurate feature detection and quantification.
• •MSFragger and IonQuant provide excellent performance for timsTOF PASEF data.
• •Flexibility allows for complex analyses, such as semi-enzymatic and open search.

     

Novel Proteome Extraction Method Illustrates a Conserved Immunological Signature of MSI-H Colorectal Tumors

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Highlights

• •TOP: robust, bio-friendly FFPE proteome extraction method with less fixation bias.
• •Proteome of MSI-H colorectal cancer identifies immunobiology key elements.
• •MSI-H tumor displays an “INFg-STAT1 centric signature”.
• •Long-term IFNg induction In-vitro mimicks MSI-H signature.

     

Phenotypic Adaption of Pseudomonas aeruginosa by Hacking Siderophores Produced by Other Microorganisms

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Highlights

• •P. aeruginosa grown with exosiderophores and analyzed by proteomic and RT-qPCR.
• •Catechol-type exosiderophores strongly induce the expression of their transporters.
• •Repression of the endogenous iron uptake pathways.
• •Complex phenotypic plasticity in the expression of the various iron-uptake pathways.

     

Seminal Plasma Proteome as an Indicator of Sperm Dysfunction and Low Sperm Motility in Chickens

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Highlights

• •Quantitative proteomes of chicken seminal plasma associated with sperm motility.
• •High abundant acrosome and mitochondrial proteins were noted in LSM seminal plasma.
• •Decreased total antioxidant capacity was highlighted in seminal plasma of LSM.
• •Lack of membrane, acrosome and mitochondrial integrity and high ROS may induce LSM.

     

An in-depth Comparison of the Pediatric and Adult Urinary N-glycomes

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Highlights

• •N-glycan patterns are distinct in pediatric and adult urine.
• •Sex differences of N-glycans are much larger in adults.
• •Pediatric urine has almost no sex differences in N-glycan levels.
• •In adults, the majority of N-glycans were more abundant in males.

     

Concentration Determination of >200 Proteins in Dried Blood Spots for Biomarker Discovery and Validation

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Highlights

• •Repeatable quantification of 200 proteins in dried blood spots.
• •Determined lower limit of quantification, repeatability, parallelism and stability.
• •Protein stability in DBS stored at ambient temperatures for up to 2 months.
• •Concentration ranges for 200 proteins in 20 healthy individuals.

     

Flow-induced Reorganization of Laminin-integrin Networks Within the Endothelial Basement Membrane Uncovered by Proteomics

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Highlights

• •Proteomics uncovers the flow-induced remodeling of the endothelial basement membrane.
• •Flow alters the composition and localization of the laminin-integrin network.
• •Flow induces proteolytic processing of LAMA4, resulting in shedding of LG4–5 region.
• •TNFα- and flow-exposure induce a distinct proteomic signature with limited interplay.

     

Molecular Dynamics Simulation-assisted Ionic Liquid Screening for Deep Coverage Proteome Analysis

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Highlights

• •Mechanistic insights into ionic liquids and proteins at molecular level.
• •Extractants prescreen for proteome analysis with MD simulation system.
• •A loss-less sample preparation method developed for in-depth proteome profiling.
• •Over 3,300 proteins were confidently identified from 1,000 HeLa cells in a 1 h run.
• •Label-free quantitative proteome analysis of human liver cancer tissues.

     

Combined EGFR and ROCK Inhibition in Triple-negative Breast Cancer Leads to Cell Death Via Impaired Autophagic Flux

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Highlights

• •We used phosphoproteomics to reveal the underlying mechanisms of drug synergy on EGFR and ROCK co-inhibition in TNBC cells.
• •EGFR inhibition alone induces autophagy activation in TNBC cells as a cytoprotective mechanism.
• •Combinatorial treatment leads to impaired autophagic flux resulting in a strong accumulation of autophagic vacuoles.
• •We hypothesize that ROCKi-induced cytoskeletal changes impair autophagosome clearance ultimately leading to cell death.

     

Organellar Maps Through Proteomic Profiling – A Conceptual Guide

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Highlights

• •Organelle profiling maps capture localizations of 1000s of proteins in one experiment.
• •Comparing maps +/− perturbation reveals disease mechanisms & cellular responses.
• •A conceptual guide to planning and interpreting organellar profiling experiments.
• •A cross-study consensus set of human organellar marker proteins.