Quantitative phosphoproteomics of transforming growth factor- Delta b signaling in colon cancer cells

• Published in: Proteomics (Weinheim) Vol. 11; no. 16; pp. 3390 - 3401
• Main Authors: Ali, Naveid A, Molloy, Mark P
• Format: Journal Article
• Language: English
• Published: 01-08-2011
• Subjects: Amino acids; Apoptosis; Casein; Cell culture; Cell division; Colon cancer; Cyclin-dependent kinase; Growth factors; Hepatoma; Isotopes; Mass spectroscopy; Nck-interacting protein; phosphoproteomes; Phosphorylation; proteomics; Signal transduction; Smad4 protein; Titanium dioxide; TRAF2 protein
• ISSN: 1615-9853; 1615-9861
• DOI: 10.1002/pmic.201100036

The transforming growth factor- Delta *b (TGF- Delta *b) signaling pathway progresses through a series of protein phosphorylation regulated steps. Smad4 is a key mediator of the classical TGF- Delta *b signaling pathway; however, reports suggest that TGF- Delta *b can activate other cellular pathways independent of Smad4. By investigating the TGF- Delta *b-regulated phosphoproteome, we aimed to uncover new functions controlled by TGF- Delta *b. We applied titanium dioxide to enrich phosphopeptides from stable isotope labeling with amino acids in cell culture (SILAC)-labeled SW480 cells stably expressing Smad4 and profiled them by mass spectrometry. TGF- Delta *b stimulation for 30min resulted in the induction of 17 phosphopeptides and the repression of 8 from a total of 149 unique phosphopeptides. Proteins previously not known to be phosphorylated by TGF- Delta *b including programmed cell death protein 4, nuclear ubiquitous casein and cyclin-dependent kinases substrate, hepatoma-derived growth factor and cell division kinases amongst others were induced following TGF- Delta *b stimulation, while the phosphorylation of TRAF2 and NCK-interacting protein kinase are examples of proteins whose phosphorylation status was repressed. This phosphoproteomic screen has identified new TGF- Delta *b-modulated phosphorylation responses in colon carcinoma cells.