Identification of an element within the promoter of human selenoprotein P responsive to transforming growth factor-beta.
Selenoprotein P (SeP) is a plasma protein that contains up to 10 selenocysteine residues and accounts for about 50% of total selenium in human plasma. We have previously shown that SeP expression in the human liver cell line HepG2 is inhibited by transforming growth factor (TGF)-beta1 on a transcriptional level. Smad proteins are the transcriptional mediators of TGF-beta signalling and putative Smad-binding elements (SBE) comprising the core sequence CAGACA are present at two positions in the SeP promoter. The aim of our study was to investigate whether Smad molecules are involved in inhibition of SeP expression by TGF-beta1 and to locate the promoter region critical for this effect. As seen in electrophoretic-mobility-shift assays, TGF-beta1 treatment led to enhanced binding of nuclear proteins to a putative SBE from the SeP promoter. Overexpression of Smad 3 and 4, but not of Smad 2, resulted in a marked down-regulation of SeP mRNA expression. Similar effects were observed for luciferase expression under control of a human SeP-promoter construct. Deletion as well as point-mutation of putative SBEs led to a loss of promoter sensitivity towards TGF-beta1 treatment. Hence, we demonstrated an involvement of Smad 3 and 4 in transcriptional regulation of SeP by TGF-beta1 and we were able to identify the TGF-beta-responsive element in the SeP promoter.
Mostert
V
V
Medizinisches Institut für Umwelthygiene an der Heinrich-Heine-Universität Düsseldorf Abteilung Experimentelle Toxikologie, Düsseldorf, Germany.
Wolff
S
S
Dreher
I
I
Köhrle
J
J
Abel
J
J
D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D004247:DNA / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D051149:Selenoprotein P; D051140:Selenoproteins; D015398:Signal Transduction; D051899:Smad2 Protein; D051900:Smad3 Protein; D051901:Smad4 Protein; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D011506:Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; C497782:SMAD2 protein, human; C497790:SMAD3 protein, human; C497796:SMAD4 protein, human; D051149:Selenoprotein P; D051140:Selenoproteins; D051899:Smad2 Protein; D051900:Smad3 Protein; D051901:Smad4 Protein; D015534:Trans-Activators; D016212:Transforming Growth Factor beta; D004247:DNA