We introduce neutron-encoded (NeuCode) amino acid labeling of mice as a strategy for multiplexed proteomic analysis in vivo. Using NeuCode, we characterize an inducible knockout mouse model of Bap1, a tumor suppressor and deubiquitinase whose in vivo roles outside of cancer are not well established. NeuCode proteomics revealed altered metabolic pathways following Bap1 deletion, including profound elevation of cholesterol biosynthetic machinery coincident with reduced expression of gluconeogenic and lipid homeostasis proteins in liver. Bap1 loss increased pancreatitis biomarkers and reduced expression of mitochondrial proteins. These alterations accompany a metabolic remodeling with hypoglycemia, hypercholesterolemia, hepatic lipid loss, and acinar cell degeneration. Liver-specific Bap1 null mice present with fully penetrant perinatal lethality, severe hypoglycemia, and hepatic lipid deficiency. This work reveals Bap1 as a metabolic regulator in liver and pancreas, and it establishes NeuCode as a reliable proteomic method for deciphering in vivo biology.
Download full-text PDF |
Source |
|---|---|
| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5546211 | PMC |
| http://dx.doi.org/10.1016/j.celrep.2016.05.096 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
mNeuCode Empowers Targeted Proteome Analysis of Arginine Dimethylation.
Anal Chem
February 2023
National Center for Protein Sciences-Beijing, Beijing Proteome Research Center, State Key Laboratory of Proteomics, Beijing Institute of Lifeomics, Beijing 102206, China.
Characterization of protein arginine dimethylation presents significant challenges due to its occurrence at the substoichiometric level. To enable a targeted MS/MS analysis of these dimethylation sites, we developed the mNeuCode (methyl-neutron-coding) tag by metabolically labeling methylarginine with stable isotopes during cell culture, which generated a diagnostic peak containing the NeuCode isotopologue signature in a high-resolution MS scan. A software tool, termed NeuCodeFinder, was developed for screening the NeuCode signatures in mass spectra.
View Article and Find Full Text PDFConstructing Human Proteoform Families Using Intact-Mass and Top-Down Proteomics with a Multi-Protease Global Post-Translational Modification Discovery Database.
J Proteome Res
October 2019
Department of Chemistry , University of Wisconsin , 1101 University Avenue, Madison , Wisconsin 53706 , United States.
Complex human biomolecular processes are made possible by the diversity of human proteoforms. Constructing proteoform families, groups of proteoforms derived from the same gene, is one way to represent this diversity. Comprehensive, high-confidence identification of human proteoforms remains a central challenge in mass spectrometry-based proteomics.
View Article and Find Full Text PDFMultiplexed proteome analysis with neutron-encoded stable isotope labeling in cells and mice.
Nat Protoc
January 2018
Genome Center, University of Wisconsin-Madison, Madison, Wisconsin, USA.
Article Synopsis
- The described protocol allows for simultaneous comparison of up to nine proteomes using NeuCode SILAC/SILAM with shorter labeling times.
- Different isotopologs of lysine are introduced to cells or mice to label the proteome based on their small mass differences.
- The entire process, including protein extraction, digestion, and mass spectrometry analysis, takes about 3-5 weeks to complete.
Proteoform Suite: Software for Constructing, Quantifying, and Visualizing Proteoform Families.
J Proteome Res
January 2018
Department of Chemistry, ‡Laboratory of Genetics, and §Genome Center of Wisconsin, University of Wisconsin-Madison, Madison, Wisconsin 53706, United States.
We present an open-source, interactive program named Proteoform Suite that uses proteoform mass and intensity measurements from complex biological samples to identify and quantify proteoforms. It constructs families of proteoforms derived from the same gene, assesses proteoform function using gene ontology (GO) analysis, and enables visualization of quantified proteoform families and their changes. It is applied here to reveal systemic proteoform variations in the yeast response to salt stress.
View Article and Find Full Text PDFElucidating Escherichia coli Proteoform Families Using Intact-Mass Proteomics and a Global PTM Discovery Database.
J Proteome Res
November 2017
Department of Chemistry, University of Wisconsin , 1101 University Avenue, Madison, Wisconsin 53706, United States.
A proteoform family is a group of related molecular forms of a protein (proteoforms) derived from the same gene. We have previously described a strategy to identify proteoforms and elucidate proteoform families in complex mixtures of intact proteins. The strategy is based upon measurements of two properties for each proteoform: (i) the accurate proteoform intact-mass, measured by liquid chromatography/mass spectrometry (LC-MS), and (ii) the number of lysine residues in each proteoform, determined using an isotopic labeling approach.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!