TNF-alpha suppresses prolyl-4-hydroxylase alpha1 expression via the ASK1-JNK-NonO pathway

Inflammation is known to contribute to the pathogenesis of vascular diseases in which arterial wall extracellular matrix (ECM) homeostasis is disrupted. Tumor necrosis factor-alpha (TNF-alpha), a pivotal cytokine that regulates ECM metabolism by increasing degradation and decreasing production of arterial collagens, is associated with vulnerable plaques and aortic aneurysms.

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.

In the current study, we showed that, when administered in doses of 1 to 100 ng/mL, TNF-alpha dose-dependently downregulated the expression of prolyl-4-hydroxylase alphaI [P4H alpha(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-alpha-responsive element (TaRE) in the human P4H alpha(I) promoter and found that a negative regulatory region at the position of -32 to +18 bp is responsible for approximately 80% of TNF-alpha-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-alpha-mediated P4H alpha 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.