Smad4/DPC4-dependent regulation of biglycan gene expression by transforming growth factor-beta in pancreatic tumor cells.
Overexpression of the small leucine-rich proteoglycan biglycan (BGN) in fibrosis and desmoplasia results from enhanced activity of transforming growth factor-beta (TGF-beta). In pancreatic adenocarcinoma, the tumor cells themselves may contribute to BGN synthesis in vivo, since 8 of 18 different pancreatic carcinoma cell lines constitutively expressed BGN mRNA, as shown by reverse transcription-PCR analysis. In PANC-1 cells, TGF-beta1 dramatically stimulated BGN mRNA accumulation through a BGN transcription-independent, cycloheximide-sensitive mechanism and strongly increased the synthesis and release of the proteoglycan form of BGN. The ability of TGF-beta1 to induce BGN mRNA was critically dependent on Smad signaling, since 1) the up-regulation of BGN mRNA was preceded by a marked increase in Smad2 phosphorylation in TGF-beta1-treated PANC-1 cells, 2) TGF-beta1 was unable to induce BGN mRNA in pancreatic carcinoma cell lines that carry homozygous deletions of the Smad4/DPC4 gene, 3) inhibition of the Smad pathway in PANC-1 cells by transfection with a dominant negative Smad4/DPC4 mutant significantly reduced TGF-beta1-induced BGN mRNA expression, 4) stable reintroduction of wild type Smad4/DPC4 into Smad4-null CFPAC-1 cells restored the TGF-beta1 effect, and 5) overexpression of Smad2 and Smad3 in PANC-1 cells augmented TGF-beta1 induction of BGN mRNA, whereas forced expression of Smad7, an inhibitory Smad, effectively blocked it. These results clearly show that a functional Smad pathway is crucial for TGF-beta regulation of BGN mRNA expression. Since BGN has been shown to inhibit growth of pancreatic cancer cells, the Smad4/DPC4 mediation of the TGF-beta effect may represent a novel tumor suppressor function for Smad4/DPC4: antiproliferation via expression of autoinhibitory BGN.
Research Unit Molecular Oncology, Clinic for General Surgery and Thoracic Surgery, Christian-Albrechts-University, D-24105 Kiel, Germany.
Wen-Bin
WB
Chen
Wolfgang
W
Lenschow
Karen
K
Tiede
Jens W
JW
Fischer
Holger
H
Kalthoff
Hendrik
H
Ungefroren
D058578:Biglycan; D002455:Cell Division; D019765:Chondroitin ABC Lyase / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D016326:Extracellular Matrix Proteins; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D005822:Genetic Vectors; D006801:Humans; D015151:Immunoblotting; D009154:Mutation; D010190:Pancreatic Neoplasms / Q000378:metabolism*; D010766:Phosphorylation; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011509:Proteoglycans / Q000378:metabolism*; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051900:Smad3 Protein; D051901:Smad4 Protein; D051906:Smad7 Protein; D015534:Trans-Activators / Q000378:metabolism*; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D015854:Up-Regulation
C546884:BGN protein, human; D058578:Biglycan; D004268:DNA-Binding Proteins; D016326:Extracellular Matrix Proteins; D011500:Protein Synthesis Inhibitors; D011509:Proteoglycans; D012333:RNA, Messenger; C497790:SMAD3 protein, human; C497796:SMAD4 protein, human; C497806:SMAD7 protein, human; D051900:Smad3 Protein; D051901:Smad4 Protein; D051906:Smad7 Protein; D015534:Trans-Activators; D016212:Transforming Growth Factor beta; D012313:RNA; D003513:Cycloheximide; D019765:Chondroitin ABC Lyase