Cellular response to hypoxia involves signaling via Smad proteins

Abstract

IntroductionThe transforming growth factor-␤ (TGF-␤) family of growth factors mediates vascular development and regulates endothelial responses to mechanical, inflammatory, and hypoxic stress. [1][2][3][4][5][6][7][8][9][10] The important role of TGF-␤ in vascular physiology is indicated by defective vasculogenesis and striking vascular inflammation leading to death in mice null for TGF-␤s, their receptors, or their downstream substrates, the Smad proteins. 3,7,11,12 We recently have shown that exposure of human umbilical vein endothelial cells (HUVECs) to hypoxia (1% O 2 ) selectively up-regulates transcription and expression of TGF-␤2 by as much as 20-fold and induces Smad2, Smad3, and Smad4 to associate with DNA. 9 In vascular endothelium, TGF-␤2, similar to TGF-␤1 and TGF-␤3, is produced in a latent form in which the bioactive, 25-kDa TGF-␤ dimer (mature TGF-␤) is noncovalently bound to its propeptide (also known as latency-associated peptide [LAP]) and is unavailable for binding to TGF-␤ membrane receptors. 1 An important physiologic regulator of TGF-␤ bioactivation is thrombospondin-1 (TSP-1), an extracellular matrix protein that is a member of the TSP family of glycoproteins. [13][14][15] TSP-1, a trimer of disulfide-linked 180-kDa subunits, is secreted from platelet ␣-granules, endothelial cells, and vascular smooth muscle cells, and is deposited in extracellular matrix. 16 Binding of TSP-1 to LAP occurs via amino acid sequence K 412 RFK 415 of TSP-1 and amino acid sequence L 54 SKL 57 of LAP, 15,17 and potentially induces a conformational change in LAP that allows interaction of the 25-kDa mature TGF-␤ peptide with its specific membrane receptors. TSP-1 can activate LAPs associated with both latent TGF-␤1 and -␤2, 15 and similarities reported between TGF-␤1-null and TSP-1-null animals 17,18 suggest that TSP-1-mediated TGF-␤ bioactivation is physiologically significant.Mature TGF-␤ can bind to its type I, type II, and type III cell membrane receptors, the first 2 of which are serine/threonine kinases. 19 Once activated by TGF-␤, the type II receptor transphosphorylates the type I receptor, which then phosphorylates Smad2 or Smad3 (receptor-activated Smads [R-Smads]), which in turn heteromerize with Smad4 (Co-Smad) to translocate to the nucleus. Smad complexes accumulate in the nucleus, where they regulate gene transcription by recruiting transcriptional coactivators or inhibitors to DNA. 19 This cross-talk created by the interplay between Smads and other signaling pathways is largely responsible for the diverse and context-specific effects of the TGF-␤ family of proteins.The Smad signaling pathway was recently shown to interact with the transacting protein complex hypoxia-inducible factor-1 (HIF-1), which is a well-characterized transcription factor complex that regulates hypoxia-driven gene expression. 20,21 HIF-1 binds DNA as a heterodimer of 2 basic helix-loop-helix proteins, HIF-1␣ and the aryl hydrocarbon receptor nuclear translocator (ARNT, or HIF-1␤). 22,23 Under normoxic conditions, HIF-1␣ is ra...

Hypoxia induces the activation of human hepatic stellate cells LX‐2 through TGF‐β signaling pathway

“…Several studies have reported that modulation of oxygen tension was recognized as an important regulator of gene activation of TGF-b in rat brain and liver, and human fibroblast and hepatoma cells [19][20][21][22]. Recently, it was also reported that cell response to hypoxia involved signaling via Smad protein in hypoxia endogelial cells by producing TGF-b with autocrine regulation [23,24]. In this study, we measured the expression level of TGF-b1 and the phosphorylation level of its downstream protein Smad2 in order to reveal the relationship between hypoxia and intracellular TGF-b signal transduction during fibrogenesis.…”

Contribution of HIF-1α in 4E-BP1 Gene Expression

“…4A). Another intriguing observation is that SMAD3/4 and HIF-1a have been shown to assemble into a same protein complex, which is responsible for transcriptional activation of different genes under hypoxia (24,25). Thus, deletion of Smad4 in BxPC-3 pancreatic cancer cells could explain the moderate induction of 4E-BP1 protein expression under hypoxia.…”

Hypoxic induction ofCtgfis directly mediated by Hif-1

“…TGF-␤ 1 activity can be increased in response to hypoxia (46,47,74); therefore, to ensure that the observed hypoxia-induced Ctgf expression was not a result of increased bioactivity of TGF-␤ 1 , we analyzed a number of luciferase promoter constructs that were rendered unresponsive to TGF-␤ 1 treatment by a previously described mutation of the SMAD binding element (25). As expected, mutation of the SMAD binding element inhibited the ability of TGF-␤ 1 to increase Ctgf promoter activity.…”