A new cellular signaling mechanism for angiotensin II activation of NF-κB -: An IκB-independent, RSK-mediated phosphorylation of p65

Objective - Angiotensin II (Ang II) promotes vascular inflammation and remodeling via activation of nuclear factor kappa B (NF-kappa B) - mediated transcription of proinflammatory genes such as interleukin-6 (IL-6). We examined the signaling mechanism whereby Ang II activates NF-kappa B in vascular smooth muscle cells (VSMCs). Methods and Results - Ang II treatment did not increase phosphorylation of inhibitor of kappa B alpha(I kappa B alpha) or I kappa B beta or decrease their levels. In contrast, mitogen-activated protein kinase kinase-1 (MEK1) inhibition (dominant-negative MEK1 adenovirus or inhibitor U0126) suppressed Ang II - induced NF-kappa B promoter activity, NF-kappa B DNA-binding activity, p65 phosphorylation, and led to 70% reduction in IL-6 transcription/production. The mechanism involved Ang II activation of Ras and MEK1. Signaling distal to MEK1 involved extracellular signal-regulated kinase (ERK) because inhibition of MEK1 suppressed the Ang II - induced activation of ribosomal S6 kinase (RSK), a substrate of ERK. Downregulation of RSK by small interfering RNA (SiRNA) in VSMCs was found to suppress Ang II - induced activation of NF-kappa B and p65 phosphorylation. Immunopurified RSK from Ang II - treated VSMCs phosphorylated recombinant glutathione S-transferase p65 in vitro. Conclusion - We uncovered a nonclassical signaling pathway (Ras/MEK1/ERK/RSK) from Ang II to activation of NF-kappa B, a mechanism by which Ang II stimulates RSK-mediated phosphorylation of p65 to participate in vascular inflammation.