Endogenous transforming growth factor-β receptor-mediated Smad signaling complexes analyzed by mass spectrometry

Qilie Luo, Edward Nieves, Julia Kzhyshkowska, Ruth Hogue Angeletti

Research output: Contribution to journalArticle

14 Scopus citations

Abstract

Smad proteins are the central feature of the transforming growth factor-β (TGF-β) intracellular signaling cascade. They function by carrying signals from the cell surface to the nucleus through through the formation of a series of signaling complexes. Changes in Smad proteins and their complexes upon treatment with TGF-β were studied in mink lung epithelial (Mv1Lu) cell cultures. A time course of incubation with TGF-β was carried out to determine the peak of appearance of phosphorylated Smad2. Immobilized monoclonal antibody against Smad2 was then used to isolate the naturally occurring complexes. Three strategies were used to identify changes in proteins partnering with Smad2: separation by one-dimensional SDS-PAGE followed by MALDI peptide mass fingerprinting, cleavable ICAT labeling of the protein mixtures analyzed by LC-MS/MS, and nano-LC followed by MALDI MS TOF/TOF. Smad2 forms complexes with many other polypeptides both in the presence and absence of TGF-β. Some of the classes of proteins identified include: transcription regulators, proteins of the cytoskeletal scaffold and other tethering proteins, motility proteins, proteins involved in transport between the cytoplasm and nucleus, and a group of membrane adaptor proteins. Although some of these have been reported in the literature, most have not been reported previously. This work expands the repertoire of proteins known to participate in the TGF-β signal transduction processes.

Original languageEnglish (US)
Pages (from-to)1245-1260
Number of pages16
JournalMolecular and Cellular Proteomics
Volume5
Issue number7
DOIs
StatePublished - Jul 1 2006

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Molecular Biology

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