TY - JOUR
T1 - Regulation of endothelial nitric-oxide synthase during hypoxia
AU - Arnet, Urs A.
AU - McMillan, Audrey
AU - Dinerman, Jay L.
AU - Ballermann, Barbara
AU - Lowenstein, Charles J.
N1 - Funding Information:
NT is a recipient of the Rio Hortega Grant (Contract 15/00246). This work was supported by grant PI15/02180 from Fondo de Investigaciones Sanitarias (Instituto de Salud Carlos III) from the Spanish Government and CIBERONC.
PY - 1996
Y1 - 1996
N2 - The mechanism by which nitric-oxide (NO) production increases during hypoxia is unknown. To explore the effect of hypoxia upon endothelial nitric- oxide synthase (ecNOS) activity and expression, we exposed bovine aortic endothelial cells (BAEC) to hypoxia (1% O2) for 0-24 h and measured levels of ecNOS mRNA, protein, and activity. The amount of ecNOS mRNA increases to more than twice the basal level after 6 h of hypoxia. Incubation of BAEC with actinomycin D during hypoxia prevents this increase, demonstrating that higher levels of mRNA observed during hypoxia are due to increased synthesis, not to increased stability of ecNOS mRNA. Levels of ecNOS protein increase throughout 24 h of hypoxia to more than twice normoxic levels. Although ecNOS expression increases within 2 h of hypoxia, total activity remains unchanged. To explore the transcriptional regulation of ecNOS, we constructed a reporter plasmid containing the ecNOS promoter region upstream of the luc gene and transfected this reporter plasmid into BAEC. In this system, hypoxia induces a linear increase over time in the expression of luciferase driven by the ecNOS promoter. It is concluded that hypoxia induces an increase in transcription of ecNOS in endothelial cells, activating the regulatory region of ecNOS by undefined transcription factors.
AB - The mechanism by which nitric-oxide (NO) production increases during hypoxia is unknown. To explore the effect of hypoxia upon endothelial nitric- oxide synthase (ecNOS) activity and expression, we exposed bovine aortic endothelial cells (BAEC) to hypoxia (1% O2) for 0-24 h and measured levels of ecNOS mRNA, protein, and activity. The amount of ecNOS mRNA increases to more than twice the basal level after 6 h of hypoxia. Incubation of BAEC with actinomycin D during hypoxia prevents this increase, demonstrating that higher levels of mRNA observed during hypoxia are due to increased synthesis, not to increased stability of ecNOS mRNA. Levels of ecNOS protein increase throughout 24 h of hypoxia to more than twice normoxic levels. Although ecNOS expression increases within 2 h of hypoxia, total activity remains unchanged. To explore the transcriptional regulation of ecNOS, we constructed a reporter plasmid containing the ecNOS promoter region upstream of the luc gene and transfected this reporter plasmid into BAEC. In this system, hypoxia induces a linear increase over time in the expression of luciferase driven by the ecNOS promoter. It is concluded that hypoxia induces an increase in transcription of ecNOS in endothelial cells, activating the regulatory region of ecNOS by undefined transcription factors.
UR - http://www.scopus.com/inward/record.url?scp=15844405458&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=15844405458&partnerID=8YFLogxK
U2 - 10.1074/jbc.271.25.15069
DO - 10.1074/jbc.271.25.15069
M3 - Article
C2 - 8663208
AN - SCOPUS:15844405458
SN - 0021-9258
VL - 271
SP - 15069
EP - 15073
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 25
ER -