Absolute Quantification of Endogenous Ras Isoform Abundance.

Abstract	Ras proteins are important signalling hubs situated near the top of networks controlling cell proliferation, differentiation and survival. Three almost identical isoforms, HRAS, KRAS and NRAS, are ubiquitously expressed yet have differing biological and oncogenic properties. In order to help understand the relative biological contributions of each isoform we have optimised a quantitative proteomics method for accurately measuring Ras isoform protein copy number per cell. The use of isotopic protein standards together with selected reaction monitoring for diagnostic peptides is sensitive, robust and suitable for application to sub-milligram quantities of lysates. We find that in a panel of isogenic SW48 colorectal cancer cells, endogenous Ras proteins are highly abundant with ≥260,000 total Ras protein copies per cell and the rank order of isoform abundance is KRAS>NRAS≥HRAS. A subset of oncogenic KRAS mutants exhibit increased total cellular Ras abundance and altered the ratio of mutant versus wild type KRAS protein. These data and methodology are significant because Ras protein copy number is required to parameterise models of signalling networks and informs interpretation of isoform-specific Ras functional data.
Authors	Craig J Mageean, John R Griffiths, Duncan L Smith, Michael J Clague, Ian A Prior
Journal	PloS one (PLoS One) Vol. 10 Issue 11 Pg. e0142674 ( 2015) ISSN: 1932-6203 [Electronic] United States
PMID	26560143 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
Chemical References	Amino Acids KRAS protein, human Membrane Proteins Protein Isoforms GTP Phosphohydrolases NRAS protein, human HRAS protein, human Proto-Oncogene Proteins p21(ras)
Topics	Amino Acids (chemistry) Cell Differentiation Cell Line, Tumor Cell Proliferation Cell Transformation, Neoplastic (genetics) Colorectal Neoplasms (metabolism) GTP Phosphohydrolases (chemistry) Gene Expression Regulation, Neoplastic Genes, ras Humans Mass Spectrometry Membrane Proteins (chemistry) Mutation Protein Isoforms (chemistry) Proteomics Proto-Oncogene Proteins p21(ras) (chemistry) Signal Transduction Tandem Mass Spectrometry

Join CureHunter, for free Research Interface BASIC access!

Take advantage of free CureHunter research engine access to explore the best drug and treatment options for any disease. Find out why thousands of doctors, pharma researchers and patient activists around the world use CureHunter every day.

Realize the full power of the drug-disease research graph!