Comparative proteomics of a model MCF10A-KRasG12V cell line reveals a distinct molecular signature of the KRasG12V cell surface.

TitleComparative proteomics of a model MCF10A-KRasG12V cell line reveals a distinct molecular signature of the KRasG12V cell surface.
Publication TypeJournal Article
Year of Publication2016
AuthorsYe, Xiaoying, Chan King C., Waters Andrew M., Bess Matthew, Harned Adam, Wei Bih-Rong, Loncarek Jadranka, Luke Brian T., Orsburn Benjamin C., Hollinger Bradley D., Stephens Robert M., Bagni Rachel, Martinko Alex, Wells James A., Nissley Dwight V., McCormick Frank, Whiteley Gordon, and Blonder Josip
JournalOncotarget
Volume7
Issue52
Pagination86948-86971
Date Published2016 Dec 27
ISSN1949-2553
Abstract

<p>Oncogenic Ras mutants play a major role in the etiology of most aggressive and deadly carcinomas in humans. In spite of continuous efforts, effective pharmacological treatments targeting oncogenic Ras isoforms have not been developed. Cell-surface proteins represent top therapeutic targets primarily due to their accessibility and susceptibility to different modes of cancer therapy. To expand the treatment options of cancers driven by oncogenic Ras, new targets need to be identified and characterized at the surface of cancer cells expressing oncogenic Ras mutants. Here, we describe a mass spectrometry-based method for molecular profiling of the cell surface using KRasG12V transfected MCF10A (MCF10A-KRasG12V) as a model cell line of constitutively activated KRas and native MCF10A cells transduced with an empty vector (EV) as control. An extensive molecular map of the KRas surface was achieved by applying, in parallel, targeted hydrazide-based cell-surface capturing technology and global shotgun membrane proteomics to identify the proteins on the KRasG12V surface. This method allowed for integrated proteomic analysis that identified more than 500 cell-surface proteins found unique or upregulated on the surface of MCF10A-KRasG12V cells. Multistep bioinformatic processing was employed to elucidate and prioritize targets for cross-validation. Scanning electron microscopy and phenotypic cancer cell assays revealed changes at the cell surface consistent with malignant epithelial-to-mesenchymal transformation secondary to KRasG12V activation. Taken together, this dataset significantly expands the map of the KRasG12V surface and uncovers potential targets involved primarily in cell motility, cellular protrusion formation, and metastasis.</p>

DOI10.18632/oncotarget.13566
Alternate JournalOncotarget
PubMed ID27894102
PubMed Central IDPMC5341332
Grant ListHHSN261200800001C / RC / CCR NIH HHS / United States
HHSN261200800001E / CA / NCI NIH HHS / United States
T32 GM064337 / GM / NIGMS NIH HHS / United States

University of Toronto  UCSF  The University of Chicago  QB3  Chicago Biomedical Consortium