TNF-α Suppresses Prolyl-4-Hydroxylase α1 Expression via the ASK1–JNK–NonO Pathway

Abstract

Background-Inflammation is known to contribute to the pathogenesis of vascular diseases in which arterial wall extracellular matrix (ECM) homeostasis is disrupted. Tumor necrosis factor-␣ (TNF-␣), a pivotal cytokine that regulates ECM metabolism by increasing degradation and decreasing production of arterial collagens, is associated with vulnerable plaques and aortic aneurysms. Methods and Results-In the current study, we showed that, when administered in doses of 1 to 100 ng/mL, TNF-␣ dose-dependently downregulated the expression of prolyl-4-hydroxylase ␣I [P4H␣(I)]-the rate-limiting subunit for the P4H enzyme essential for procollagen hydroxylation, secretion, and deposition in primary human aortic smooth muscle cells (HASMCs). Using a progressive deletion cloning approach, we characterized the TNF-␣-responsive element (TaRE) in the human P4H␣(I) promoter and found that a negative regulatory region at the position of Ϫ32 to ϩ18bp is responsible for Ϸ80% of TNF-␣-mediated suppression. Using oligonucleotide-based transcription factor pull-down method in which proteins were resolved in 1-D gel electrophoresis and identified using LC-MS/MS, we identified the NonO protein binds this region. When NonO expression silenced with specific siRNA, we found that 70% of the TNF-␣-mediated P4H␣ suppression was abolished, which appeared to be mediated by the ASK1-JNK pathway. Conclusions-Our findings define a novel molecular pathway for inflammation associated extracellular matrix dysregulation, which may account for atherosclerotic plaque rupture and aortic aneurysm formation. Further understanding of this pathway may facilitate development of novel therapeutics for vascular diseases. (Arterioscler Thromb Vasc Biol.

Effect of Adiponectin Overexpression on Stability of Preexisting Plaques by Inducing Prolyl-4-Hydroxylase Expression

“…28-30 Several studies have revealed that various cytokines, such as interleukins and TNF-α, inhibit the P4H isoenzymes. Zhang et al reported that TNF-α suppresses collagen synthesis by specifically inhibiting P4Hα1, 31 and Raveendran et al also showed that cigarette smoke extract can significantly downregulate the expression of P4Hα1. 32 However, little information exists on whether the defense molecules regulating the expression of P4Hα1 play a key role in plaque cap stability.…”

Effect of 27nt Small RNA on Endothelial Nitric-Oxide Synthase Expression

“…Acetylation of the histones resulted in recruitment of the NonO protein, which has been shown to have a suppressive effect on gene transcription (Zhang et al, 2007). Through protein-protein interaction, nuclear actin, which binds on the 27nt repeat element, recruits HDAC3 to the site.…”

“…We further detected interaction between HDAC3 and NonO protein ( Figure 3B, middle left), and between nuclear actin and NonO ( Figure 3B, bottom left). Our previous work showed that NonO protein, when recruited to the target genomic location, is frequently associated with suppression of transcription (Zhang et al, 2007). The recruitment of the NonO protein to the HDAC3 and nuclear actin complex in the 27nt repeat region could be responsible for the suppressive effects of the 27nt small RNA on eNOS transcription.…”

Role of Adiponectin in Cardiovascular Protection

“…11 P4H expression is also regulated by NO, TNF-α, transforming growth factor-β, insulinlike growth factor-1, β fibroblast growth factor, cytokines, and even cigarette smoking as well. 12 It has been reported that the anti-atherogenic effect of adiponectin was in part mediated by its regulatory ability on collagen synthesis, 13 but precise molecular mechanism was unknown. In the current study by Li and co-workers, they newly identified that adiponectin is another player for the regulation of P4H expression.…”