Smad3 is a key nonredundant mediator of transforming growth factor beta signaling in Nme mouse mammary epithelial cells.
Smad2 and Smad3 are intracellular mediators of transforming growth factor beta (TGFbeta) signaling that share various biochemical properties, but data emerging from functional analyses in several cell types indicate that these two Smad proteins may convey distinct cellular responses. Therefore, we have investigated the individual roles of Smad2 and Smad3 in mediating the cytostatic and proapoptotic effects of TGFbeta as well as their function in epithelial-to-mesenchymal transition. For this purpose, we transiently depleted mouse mammary epithelial cells (Nme) of Smad2 and/or Smad3 mainly by a strategy relying on RNaseH-induced degradation of mRNA. The effect of such depletion on hallmark events of TGFbeta-driven epithelial-to-mesenchymal transition was analyzed, including dissolution of epithelial junctions, formation of stress fibers and focal adhesions, activation of metalloproteinases, and transcriptional regulation of acknowledged target genes. Furthermore, we investigated the effect of Smad2 and Smad3 knockdown on the TGFbeta-regulated transcriptome by microarray analysis. Our results identify Smad3 as a key factor to trigger TGFbeta-regulated events and ascribe tumor suppressor as well as oncogenic activities to this protein.
Dzwonek
Joanna
J
Laboratory of Molecular Biology (Celgen), Center for Human Genetics, Campus Gasthuisberg K.U.Leuven, Leuven, Belgium.
Preobrazhenska
Olena
O
Cazzola
Silvia
S
Conidi
Andrea
A
Schellens
Ann
A
van Dinther
Maarten
M
Stubbs
Andrew
A
Klippel
Anke
A
Huylebroeck
Danny
D
ten Dijke
Peter
P
Verschueren
Kristin
K
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D048430:Cell Shape / Q000187:drug effects; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D004790:Enzyme Induction / Q000187:drug effects; D004847:Epithelial Cells / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D005260:Female; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects; D055785:Gene Knockdown Techniques; D006801:Humans; D007365:Intercellular Junctions / Q000187:drug effects / Q000378:metabolism; D008321:Mammary Glands, Animal / Q000166:cytology*; D020782:Matrix Metalloproteinases / Q000096:biosynthesis; D051379:Mice; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D051899:Smad2 Protein / Q000235:genetics / Q000378:metabolism; D051900:Smad3 Protein / Q000235:genetics / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000378:metabolism*
D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D051899:Smad2 Protein; C497783:Smad2 protein, mouse; D051900:Smad3 Protein; C497791:Smad3 protein, mouse; D016212:Transforming Growth Factor beta; D020782:Matrix Metalloproteinases