Endoglin increases eNOS expression by modulating Smad2 protein levels and Smad2-dependent TGF-beta signaling.
The endothelial nitric oxide synthase (eNOS) is a critical regulator of cardiovascular homeostasis, whose dysregulation leads to different vascular pathologies. Endoglin is a component of the transforming growth factor beta (TGF-beta) receptor complex present in endothelial cells that is involved in angiogenesis, cardiovascular development, and vascular homeostasis. Haploinsufficient expression of endoglin has been shown to downregulate endothelium-derived nitric oxide in endoglin(+/-) (Eng(+/-)) mice and cultured endothelial cells. Here, we find that TGF-beta1 leads to an increased vasodilatation in Eng(+/+) mice that is severely impaired in Eng(+/-) mice, suggesting the involvement of endoglin in the TGF-beta regulated vascular homeostasis. The endoglin-dependent induction of eNOS occurs at the transcriptional level and is mediated by the type I TGF-beta receptor ALK5 and its downstream substrate Smad2. In addition, Smad2-specific signaling is upregulated in endoglin-induced endothelial cells, whereas it is downregulated upon endoglin gene suppression with small interference RNA (siRNA). The endoglin-dependent upregulation of Smad2 was confirmed using eNOS and pARE promoters, whose activities are known to be Smad2 dependent, as well as with the interference of Smad2 with siRNA, Smurf2, or a dominant negative form of Smad2. Furthermore, increased expression of endoglin in endoglin-inducible endothelial cells or in transfectants resulted in increased levels of Smad2 protein without affecting the levels of Smad2 mRNA. The increased levels of Smad2 appear to be due to a decreased ubiquitination and proteasome-dependent degradation leading to stabilization of Smad2. These results suggest that endoglin enhances Smad2 protein levels potentiating TGF-beta signaling, and leading to an increased eNOS expression in endothelial cells.
Santibanez
Juan F
JF
Centro de Investigaciones Biologicas, CSIC, Madrid, Spain.
Letamendia
Ainhoa
A
Perez-Barriocanal
Fernando
F
Silvestri
Cristoforo
C
Saura
Marta
M
Vary
Calvin P H
CP
Lopez-Novoa
Jose M
JM
Attisano
Liliana
L
Bernabeu
Carmelo
C
D000818:Animals; D019556:COS Cells; D002417:Cattle; D002522:Chlorocebus aethiops; D000071063:Endoglin; D042783:Endothelial Cells / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D006706:Homeostasis / Q000502:physiology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D009569:Nitric Oxide / Q000096:biosynthesis; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction / Q000502:physiology*; D051899:Smad2 Protein / Q000235:genetics / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D015854:Up-Regulation / Q000235:genetics*; D014664:Vasodilation / Q000235:genetics
D000071063:Endoglin; C502004:Eng protein, mouse; D047908:Intracellular Signaling Peptides and Proteins; D012333:RNA, Messenger; D051899:Smad2 Protein; C497783:Smad2 protein, mouse; D016212:Transforming Growth Factor beta; D009569:Nitric Oxide; D052250:Nitric Oxide Synthase Type III