RRC ID 67346
Author Kauko O, Laajala TD, Jumppanen M, Hintsanen P, Suni V, Haapaniemi P, Corthals G, Aittokallio T, Westermarck J, Imanishi SY.
Title Label-free quantitative phosphoproteomics with novel pairwise abundance normalization reveals synergistic RAS and CIP2A signaling.
Journal Sci Rep
Abstract Hyperactivated RAS drives progression of many human malignancies. However, oncogenic activity of RAS is dependent on simultaneous inactivation of protein phosphatase 2A (PP2A) activity. Although PP2A is known to regulate some of the RAS effector pathways, it has not been systematically assessed how these proteins functionally interact. Here we have analyzed phosphoproteomes regulated by either RAS or PP2A, by phosphopeptide enrichment followed by mass-spectrometry-based label-free quantification. To allow data normalization in situations where depletion of RAS or PP2A inhibitor CIP2A causes a large uni-directional change in the phosphopeptide abundance, we developed a novel normalization strategy, named pairwise normalization. This normalization is based on adjusting phosphopeptide abundances measured before and after the enrichment. The superior performance of the pairwise normalization was verified by various independent methods. Additionally, we demonstrate how the selected normalization method influences the downstream analyses and interpretation of pathway activities. Consequently, bioinformatics analysis of RAS and CIP2A regulated phosphoproteomes revealed a significant overlap in their functional pathways. This is most likely biologically meaningful as we observed a synergistic survival effect between CIP2A and RAS expression as well as KRAS activating mutations in TCGA pan-cancer data set, and synergistic relationship between CIP2A and KRAS depletion in colony growth assays.
Volume 5
Pages 13099
Published 2015-8-17
DOI 10.1038/srep13099
PII srep13099
PMID 26278961
PMC PMC4642524
MeSH Area Under Curve Autoantigens / genetics Autoantigens / metabolism* Cell Proliferation Chromatography, High Pressure Liquid Cluster Analysis HeLa Cells Humans Intracellular Signaling Peptides and Proteins Kaplan-Meier Estimate Membrane Proteins / antagonists & inhibitors Membrane Proteins / genetics Membrane Proteins / metabolism* Neoplasms / metabolism Neoplasms / mortality Neoplasms / pathology Phosphopeptides / analysis* Phosphorylation Protein Phosphatase 2 / metabolism RNA Interference RNA, Small Interfering / metabolism ROC Curve Signal Transduction Tandem Mass Spectrometry Titanium / chemistry ras Proteins / antagonists & inhibitors ras Proteins / genetics ras Proteins / metabolism*
IF 3.998
Human and Animal Cells CW-2(RCB0778)