title: Proteomic analysis of the TGF-beta signaling pathway in pancreatic carcinoma cells using stable RNA interference to silence Smad4 expression.
abstract: Smad4 is a tumor-suppressor gene that is lost or mutated in 50% of pancreatic carcinomas. Smad4 is also an intracellular transmitter of transforming growth factor-beta (TGF-beta) signals. Although its tumor-suppressor function is presumed to reside in its capacity to mediate TGF-beta-induced growth inhibition, there seems to be a Smad4-independent TGF-beta signaling pathway. Here, we succeeded in establishing Smad4 knockdown (S4KD) pancreatic cancer cell lines using stable RNA interference. Smad4 protein expression and TGF-beta-Smad4 signaling were impaired in S4KD cells, and we compared the proteomic changes with TGF-beta stimulation using two-dimensional gel electrophoresis (2-DE) and mass spectrometry. We identified five proteins that were up-regulated and seven proteins that were down-regulated; 10 of them were novel targets for TGF-beta. These proteins function in processes such as cytoskeletal regulation, cell cycle, and oxidative stress. Introducing siRNA-mediated gene silencing into proteomics revealed a novel TGF-beta signal pathway that did not involve Smad4.
authors.affiliation: Department of Gastroenterology, Graduate School of Medicine, University of Tokyo, Tokyo, Japan.
authors.forename: Takaaki
authors.initials: T
authors.lastname: Imamura
authors.forename: Fumihiko
authors.initials: F
authors.lastname: Kanai
authors.forename: Takayuki
authors.initials: T
authors.lastname: Kawakami
authors.forename: Jazag
authors.initials: J
authors.lastname: Amarsanaa
authors.forename: Hideaki
authors.initials: H
authors.lastname: Ijichi
authors.forename: Yujin
authors.initials: Y
authors.lastname: Hoshida
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Tanaka
authors.forename: Tsuneo
authors.initials: T
authors.lastname: Ikenoue
authors.forename: Keisuke
authors.initials: K
authors.lastname: Tateishi
authors.forename: Takao
authors.initials: T
authors.lastname: Kawabe
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Arakawa
authors.forename: Makoto
authors.initials: M
authors.lastname: Miyagishi
authors.forename: Kazunari
authors.initials: K
authors.lastname: Taira
authors.forename: Osamu
authors.initials: O
authors.lastname: Yokosuka
authors.forename: Masao
authors.initials: M
authors.lastname: Omata
mesh_terms: D015153:Blotting, Western; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics*; D015180:Electrophoresis, Gel, Two-Dimensional; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D020868:Gene Silencing; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D010190:Pancreatic Neoplasms / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D014176:Protein Biosynthesis; D011506:Proteins / Q000032:analysis; D020543:Proteome / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference / Q000502:physiology*; D015398:Signal Transduction; D051901:Smad4 Protein; D015534:Trans-Activators / Q000096:biosynthesis* / Q000235:genetics*; D016212:Transforming Growth Factor beta / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D005419:Flavonoids; D011506:Proteins; D020543:Proteome; C497796:SMAD4 protein, human; D051901:Smad4 Protein; D015534:Trans-Activators; D016212:Transforming Growth Factor beta; D020928:Mitogen-Activated Protein Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one