Dynamic ADP-Ribosylome, Phosphoproteome, and Interactome in LPS-Activated Macrophages.
Casey M DanielsPauline R KaplanIsaac BishofClinton BradfieldTrisha TucholskiArthur G NuccioNathan P ManesSamuel KatzIain D C FraserAleksandra Nita-LazarPublished in: Journal of proteome research (2020)
We have used mass spectrometry (MS) to characterize protein signaling in lipopolysaccharide (LPS)-stimulated macrophages from human blood, human THP1 cells, mouse bone marrow, and mouse Raw264.7 cells. Protein ADP-ribosylation was truncated down to phosphoribose, allowing for enrichment and identification of the resulting phosphoribosylated peptides alongside phosphopeptides. Size exclusion chromatography-MS (SEC-MS) was used to separate proteoforms by size; protein complexes were then identified by weighted correlation network analysis (WGCNA) based on their correlated movement into or out of SEC fractions following stimulation, presenting an analysis method for SEC-MS that does not rely on established databases. We highlight two modules of interest: one linked to the apoptosis signal-regulating kinase (ASK) signalosome and the other containing poly(ADP-ribose) polymerase 9 (PARP9). Finally, PARP inhibition was used to perturb the characterized systems, demonstrating the importance of ADP-ribosylation for the global interactome. All post-translational modification (PTM) and interactome data have been aggregated into a meta-database of 6729 proteins, with ADP-ribosylation characterized on 2905 proteins and phosphorylation characterized on 2669 proteins. This database-titled MAPCD, for Macrophage ADP-ribosylation, Phosphorylation, and Complex Dynamics-serves as an invaluable resource for studying crosstalk between the ADP-ribosylome, phosphoproteome, and interactome.
Keyphrases
- mass spectrometry
- network analysis
- cell cycle arrest
- liquid chromatography
- induced apoptosis
- multiple sclerosis
- ms ms
- bone marrow
- endothelial cells
- inflammatory response
- gas chromatography
- amino acid
- oxidative stress
- high performance liquid chromatography
- cell death
- protein protein
- protein kinase
- high resolution
- magnetic resonance
- dna repair
- tandem mass spectrometry
- emergency department
- binding protein
- adipose tissue
- big data
- computed tomography
- electronic health record
- case report
- drug induced
- data analysis