Hypoxia up-regulates hypoxia-inducible factor-1alpha expression through RhoA activation in trophoblast cells.
During early pregnancy, trophoblast cells are exposed to relatively low-oxygen tension. Recently, the Rho GTPase family has been shown to play a key role in hypoxia-inducible factor-1 (HIF-1) alpha induction in renal cell carcinoma. The present study was designed to investigate the effect of low-oxygen conditions on RhoA expression in trophoblast cells isolated from early stages of human placenta and in trophoblast-derived BeWo cells and JAR cells. Immunoblot and RT-PCR analyses showed that low-oxygen conditions (1% O(2) or 250 mum CoCl(2)) stimulated expression of RhoA protein and mRNA. Pull-down assays demonstrated that these low-oxygen conditions increased RhoA activity. Preincubation of BeWo cells with Clostridium botulinum C3 exoenzyme, a specific inhibitor of Rho, inhibited hypoxia-induced HIF-1alpha expression. Under 1% O(2) or 250 mum CoCl(2), BeWo cells, transfected with a dominant-negative RhoA, exhibited decreased levels of HIF-1alpha protein and mRNA compared with the control vector transfectants. BeWo cells expressing constitutively active RhoA showed enhanced protein levels of not only HIF-1alpha but also vascular endothelial growth factor (VEGF) and glucose transporter 1, which are target gene products of HIF-1alpha. These findings suggest that up-regulation of RhoA induced by low-oxygen conditions may play an important role in regulation of HIF-1alpha expression in trophoblast cells.
Hayashi
Masami
M
Department of Obstetrics and Gynecology, Osaka University, Faculty of Medicine, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
Sakata
Masahiro
M
Takeda
Takashi
T
Tahara
Masahiro
M
Yamamoto
Toshiya
T
Minekawa
Ryoko
R
Isobe
Aki
A
Tasaka
Keiichi
K
Murata
Yuji
Y
D036002:ADP Ribose Transferases / Q000494:pharmacology; D001905:Botulinum Toxins / Q000494:pharmacology; D002478:Cells, Cultured; D005260:Female; D015870:Gene Expression / Q000502:physiology; D051272:Glucose Transporter Type 1; D006801:Humans; D000860:Hypoxia / Q000378:metabolism / Q000503:physiopathology*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009004:Monosaccharide Transport Proteins / Q000235:genetics; D010100:Oxygen / Q000494:pharmacology; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014327:Trophoblasts / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics; D020742:rhoA GTP-Binding Protein / Q000037:antagonists & inhibitors / Q000378:metabolism*
D051272:Glucose Transporter Type 1; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009004:Monosaccharide Transport Proteins; D012333:RNA, Messenger; C495677:SLC2A1 protein, human; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; D001905:Botulinum Toxins; D020742:rhoA GTP-Binding Protein; D010100:Oxygen