Journal article
Global view of the RAF-MEK-ERK module and its immediate downstream effectors
Department of Biotechnology and Biomedicine, Technical University of Denmark1
University of Copenhagen2
DTU Proteomics Core, Section for Protein Science and Biotherapeutics, Department of Biotechnology and Biomedicine, Technical University of Denmark3
Yale University4
Massachusetts Institute of Technology5
Samsung Genome Institute6
Icahn School of Medicine at Mount Sinai7
Memorial Sloan-Kettering Cancer Center8
Small molecule inhibitors of BRAF and MEK have proven effective at inhibiting tumor growth in melanoma patients, however this efficacy is limited due to the almost universal development of drug resistance. To provide advanced insight into the signaling responses that occur following kinase inhibition we have performed quantitative (phospho)-proteomics of human melanoma cells treated with either dabrafenib, a BRAF inhibitor; trametinib, a MEK inhibitor or SCH772984, an ERK inhibitor.
Over nine experiments we identified 7827 class I phosphorylation sites on 4960 proteins. This included 54 phosphorylation sites that were significantly down-modulated after exposure to all three inhibitors, 34 of which have not been previously reported. Functional analysis of these novel ERK targets identified roles for them in GTPase activity and regulation, apoptosis and cell-cell adhesion.
Comparison of the results presented here with previously reported phosphorylation sites downstream of ERK showed a limited degree of overlap suggesting that ERK signaling responses may be highly cell line and cue specific. In addition we identified 26 phosphorylation sites that were only responsive to dabrafenib.
We provide further orthogonal experimental evidence for 3 of these sites in human embryonic kidney cells over-expressing BRAF as well as further computational insights using KinomeXplorer. The validated phosphorylation sites were found to be involved in actin regulation, which has been proposed as a novel mechanism for inhibiting resistance development.
These results would suggest that the linearity of the BRAF-MEK-ERK module is at least context dependent.
Language: | English |
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Publisher: | Nature Publishing Group UK |
Year: | 2019 |
Pages: | 10865 |
ISSN: | 20452322 |
Types: | Journal article |
DOI: | 10.1038/s41598-019-47245-x |
ORCIDs: | 0000-0002-8645-2707 , 0000-0003-4535-5000 , 0000-0001-6271-7750 , 0000-0001-9524-9981 and Schoof, Erwin M. |
Apoptosis BRAF protein, human Cancer Cell Line, Tumor Cell signalling Drug Resistance, Neoplasm Extracellular Signal-Regulated MAP Kinases Humans Imidazoles Indazoles MAP Kinase Signaling System Mass spectrometry Melanoma Mitogen-Activated Protein Kinase Kinases Oximes Phosphorylation Piperazines Proteome Proteomics Proto-Oncogene Proteins B-raf Pyridones Pyrimidinones SCH772984 Skin Neoplasms dabrafenib trametinib