title: TGF-beta-induced nuclear localization of Smad2 and Smad3 in Smad4 null cancer cell lines.
abstract: Smad4 is a tumor suppressor gene that is commonly lost or mutated in colorectal and pancreatic cancers. The activated transforming growth factor-beta (TGF-beta) receptor phosphorylates Smad2 and Smad3, which then complex with Smad4 and translocate to the nucleus. Smad4 mutations when detected as present in some human cancers have been considered sufficient to inactivate TGF-beta signaling. In this work, we describe a colon cancer cell line, VACO-9M, that is Smad4 null when analysed by multiple assays. To study the role of Smad4 in TGF-beta-induced translocation of the receptor-activated Smads to the nucleus, we analysed by immunofluorescence the cellular localization of endogenous Smad2 and Smad3 after TGF-beta treatment of VACO-9M, plus four additional Smad4 null cell lines of breast (MDA-MB-468), or pancreatic (BxPC3, Hs766T, CFPAC-1) origin. In each cell line, TGF-beta treatment resulted in both Smad2 and Smad3 moving to the nucleus in a Smad4-independent fashion. Nuclear translocation of Smad2 and Smad3 was, however, not sufficient to activate reporters for TGF-beta-induced transcriptional responses, which were however restored by transient transfection of wild-type Smad4. We conclude that Smad4 is not required for nuclear translocation of Smad2 and Smad3, but is needed for activation of at least certain transcriptional responses.
authors.affiliation: Howard Hughes Medical Institute, Cleveland, OH 44106, USA.
authors.forename: Stephen P
authors.initials: SP
authors.lastname: Fink
authors.forename: Debra
authors.initials: D
authors.lastname: Mikkola
authors.forename: James K V
authors.initials: JK
authors.lastname: Willson
authors.forename: Sanford
authors.initials: S
authors.lastname: Markowitz
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects*; D000230:Adenocarcinoma / Q000473:pathology; D001943:Breast Neoplasms / Q000473:pathology; D002467:Cell Nucleus / Q000378:metabolism; D003110:Colonic Neoplasms / Q000473:pathology; D004268:DNA-Binding Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D017930:Genes, Reporter; D006801:Humans; D008856:Microscopy, Fluorescence; D009363:Neoplasm Proteins / Q000378:metabolism*; D010190:Pancreatic Neoplasms / Q000473:pathology; D011993:Recombinant Fusion Proteins / Q000502:physiology; D051899:Smad2 Protein; D051900:Smad3 Protein; D051901:Smad4 Protein; D015534:Trans-Activators / Q000172:deficiency / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D014407:Tumor Cells, Cultured / Q000187:drug effects
chemical_list: D004268:DNA-Binding Proteins; D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins; C497782:SMAD2 protein, human; C497790:SMAD3 protein, human; C497796:SMAD4 protein, human; D051899:Smad2 Protein; D051900:Smad3 Protein; D051901:Smad4 Protein; D015534:Trans-Activators; D016212:Transforming Growth Factor beta