The PAI-1 gene as a direct target of endothelial PAS domain protein-1 in adenocarcinoma A549 cells.
Endothelial PAS domain protein-1 (EPAS1) regulates transcription of the genes encoding erythropoietin and vascular endothelial growth factor, which are important for maintaining oxygen homeostasis. We have previously shown that plasminogen activator inhibitor-1 (PAI-1) gene expression is induced by hypoxia. In this study, we sought to determine whether PAI-1 gene expression is directly regulated by EPAS1 in cancer cells because activities of proteases and their inhibitors are tightly regulated for tumor invasion. Hypoxia increased the PAI-1 mRNA levels in human adenocarcinoma A549 cells. Overexpression of EPAS1 significantly increased the PAI-1 mRNA and protein levels. Transient transfection assays revealed that EPAS1 increased PAI-1 gene transcription through a sequence containing 5'-CACGTACA-3' located at -194 (we refer to it as site HREPAI-1) and GT-box located at -78. Electrophoretic gel mobility shift assays revealed that HREPAI-1 serves as a binding site for EPAS1, and Sp1 constitutively binds to GT-box. In conclusion, PAI-1 expression is induced by EPAS1 through HREPAI-1 and through an Sp1-binding site. These results indicate that the PAI-1 gene is a direct target of EPAS1 and suggest the role of EPAS1 and Sp1 in the hypoxic response of cancer cells.
Sato
Mahito
M
Second Department of Internal Medicine, Gunma University School of Medicine, Japan.
Tanaka
Toru
T
Maemura
Koji
K
Uchiyama
Tsuyoshi
T
Sato
Hiroko
H
Maeno
Toshitaka
T
Suga
Tatsuo
T
Iso
Tatsuya
T
Ohyama
Yoshio
Y
Arai
Masashi
M
Tamura
Junichi
J
Sakamoto
Hironosuke
H
Nagai
Ryozo
R
Kurabayashi
Masahiko
M
D000230:Adenocarcinoma / Q000378:metabolism* / Q000473:pathology; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins; D045744:Cell Line, Tumor; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011506:Proteins / Q000235:genetics*; D016329:Sp1 Transcription Factor / Q000378:metabolism / Q000502:physiology; D051705:Sp3 Transcription Factor; D014157:Transcription Factors / Q000378:metabolism; D000099162:Endothelial PAS Domain-Containing Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D017346:Protein Serine-Threonine Kinases; D011506:Proteins; D016329:Sp1 Transcription Factor; D051705:Sp3 Transcription Factor; D014157:Transcription Factors; D000099162:Endothelial PAS Domain-Containing Protein 1; C116037:PAS domain kinases