title: Quantitative Proteomics of the SMAD (Suppressor of Mothers against Decapentaplegic) Transcription Factor Family Identifies Importin 5 as a Bone Morphogenic Protein Receptor SMAD-specific Importin.
abstract: Gene-specific transcription factors (GSTFs) control gene transcription by DNA binding and specific protein complex recruitment, which regulates promoter accessibility for transcription initiation by RNA polymerase II. Mutations in the GSTFs Suppressor of Mothers Against Decapentaplegic 2 (SMAD2) and SMAD4 are frequently associated with colon and rectal carcinomas. These proteins play an important role in bone morphogenic protein (BMP) and transforming growth factor β (TGF-β) signaling pathways controlling cell fate and proliferation. To study the protein interactome of the SMAD protein family we generated a quantitative proteomics pipeline that allows for inducible expression of GFP-tagged SMAD proteins followed by affinity purification and quantitative mass spectrometry analysis. Data are available via ProteomeXchange with identifier PXD004529. The nuclear importin IPO5 was identified as a novel interacting protein of SMAD1. Overexpression of IPO5 in various cell lines specifically increases nuclear localization of BMP receptor-activated SMADs (R-SMADs) confirming a functional relationship between IPO5 and BMP but not TGF-β R-SMADs. Finally, we provide evidence that variation in length of the lysine stretch of the nuclear localization sequence is a determinant for importin specificity.
authors.affiliation: From the Departments of Molecular Cancer Research and Stem Cells, Regenerative Medicine Center, Center for Molecular Medicine, University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands.
authors.forename: Roy
authors.initials: R
authors.lastname: Baas
authors.affiliation: From the Departments of Molecular Cancer Research and Stem Cells, Regenerative Medicine Center, Center for Molecular Medicine, University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands.
authors.forename: Ayestha
authors.initials: A
authors.lastname: Sijm
authors.affiliation: From the Departments of Molecular Cancer Research and Stem Cells, Regenerative Medicine Center, Center for Molecular Medicine, University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands.
authors.forename: Hetty A A M
authors.initials: HA
authors.lastname: van Teeffelen
authors.affiliation: From the Departments of Molecular Cancer Research and Stem Cells, Regenerative Medicine Center, Center for Molecular Medicine, University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands.
authors.forename: Robert
authors.initials: R
authors.lastname: van Es
authors.affiliation: From the Departments of Molecular Cancer Research and Stem Cells, Regenerative Medicine Center, Center for Molecular Medicine, University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands.
authors.forename: Harmjan R
authors.initials: HR
authors.lastname: Vos
authors.affiliation: From the Departments of Molecular Cancer Research and Stem Cells, Regenerative Medicine Center, Center for Molecular Medicine, University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands H.T.M.Timmers@umcutrecht.nl.
authors.forename: H Th
authors.initials: HT
authors.lastname: Marc Timmers
mesh_terms: D052004:Bone Morphogenetic Protein Receptors / Q000235:genetics / Q000378:metabolism; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D040901:Proteomics; D051898:Smad1 Protein / Q000235:genetics / Q000378:metabolism*; D051899:Smad2 Protein / Q000235:genetics / Q000378:metabolism*; D051901:Smad4 Protein / Q000235:genetics / Q000378:metabolism*; D028961:beta Karyopherins / Q000235:genetics / Q000378:metabolism*
chemical_list: C107940:IPO5 protein, human; C497399:SMAD1 protein, human; C497782:SMAD2 protein, human; C497796:SMAD4 protein, human; D051898:Smad1 Protein; D051899:Smad2 Protein; D051901:Smad4 Protein; D028961:beta Karyopherins; D052004:Bone Morphogenetic Protein Receptors
keywords: SMAD transcription factor; mass spectrometry (MS); nuclear translocation; proteomics; transcription regulation