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Mol Cell. 2009 Apr 10;34(1):115-31. doi: 10.1016/j.molcel.2009.03.007.

Functional proteomics identifies targets of phosphorylation by B-Raf signaling in melanoma.

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1
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO 80309-0215, USA.

Abstract

Melanoma and other cancers harbor oncogenic mutations in the protein kinase B-Raf, which leads to constitutive activation and dysregulation of MAP kinase signaling. In order to elucidate molecular determinants responsible for B-Raf control of cancer phenotypes, we present a method for phosphoprotein profiling, using negative ionization mass spectrometry to detect phosphopeptides based on their fragment ion signature caused by release of PO(3)(-). The method provides an alternative strategy for phosphoproteomics, circumventing affinity enrichment of phosphopeptides and isotopic labeling of samples. Ninety phosphorylation events were regulated by oncogenic B-Raf signaling, based on their responses to treating melanoma cells with MKK1/2 inhibitor. Regulated phosphoproteins included known signaling effectors and cytoskeletal regulators. We investigated MINERVA/FAM129B, a target belonging to a protein family with unknown category and function, and established the importance of this protein and its MAP kinase-dependent phosphorylation in controlling melanoma cell invasion into three-dimensional collagen matrix.

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PMID:
19362540
PMCID:
PMC2735263
DOI:
10.1016/j.molcel.2009.03.007
[Indexed for MEDLINE]
Free PMC Article

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