Abstract 175: The Nck Adaptor Proteins Couple Oxidant Stress-Driven Tyrosine Phosphorylation to NF-κB Activation and Proinflammatory Gene Expression
Reactive oxygen species production drives endothelial NF-kappaB activation and NF-kappaB-dependent proinflammatory gene expression in several pathological conditions, including ischemia-reperfusion injury. While multiple reports link oxidant stress-induced tyrosine kinase signaling to NF-kappaB, we previously showed that the serine/threonine kinase p21 activated kinase (PAK) critically mediates NF-kappaB activation by oxidant stress. The adaptor protein Nck classically couples PAK to tyrosine kinase signaling by recruiting PAK to the plasma membrane. However, the mechanisms linking oxidant stress to PAK signaling are currently unknown. We now show that H2O2 stimulates both Nck interactions with tyrosine phosphorylated proteins and membrane translocation, whereas inhibiting Nck (siRNA, blocking peptide) abrogates H2O2-induced PAK activation. Consistent with PAK’s role in NF-kappaB signaling, Nck knockdown completely blocks H2O2-induced NF-kappaB activation, proinflammatory gene expression, and monocyte adhesion, suggesting that Nck couples tyrosine kinase signaling to PAK-dependent NF-kappaB activation. In contrast to these proinflammatory responses, antioxidant gene expression following oxidant stress is not affected by Nck inhibition. In addition to exogenous H2O2, endogenously produced oxidant stress following hypoxia/reoxygenation injury shows a similar requirement for Nck for proinflammatory signaling (PAK, NF-kappaB), proinflammatory gene expression (ICAM-1, VCAM-1) and monocyte adhesion. Finally, leukocyte firm adhesion and emigration following ischemia-reperfusion injury in the cremaster muscle of C57Bl/6J mice is completely inhibited by treatment with the Nck blocking peptide. Taken together, these data identify Nck as an important mediator of oxidant stress-induced inflammation and a potential therapeutic target for ischemia/reperfusion injury.
Author Disclosures: J. Chen: None. I. Leskov: None. P. Albert: None. C.G. Kevil: None. K. Stokes: None. A.W. Orr: None.
This research has received full or partial funding support from the American Heart Association.
- © 2014 by American Heart Association, Inc.