Identification of Signal-induced IκB-α Kinases in Human Endothelial Cells

Abstract

Activation of the nuclear transcription factor-B is an early event in endothelial activation. NF-B activation is regulated by the inducible phosphorylation and subsequent degradation of the inhibitory subunit IB-␣. We identified two discrete kinases of approximately 36 and 41 kDa in the cytoplasm of human umbilical vein endothelial cells that specifically bind to and phosphorylate the IB-␣ subunit. IB-␣ kinase activity is transiently elevated following treatment with either tumor necrosis factor ␣, interleukin-1␤, or bacterial lipopolysaccharides and precedes activation of either mitogen-activated kinase or Jun kinase. Furthermore, activation of the IB-␣ kinases precedes both the appearance of hyperphosphorylated IB-␣ and its subsequent degradation, as well as the translocation of NF-B to the nucleus. Deletion mutagenesis of the IB-␣ polypeptide revealed that these kinases bind in or around the ankyrin repeat domains and phosphorylate residues within the C terminus. These kinases, however, were not identical to casein kinase II and displayed a pharmacologic profile distinct from other known kinases. These kinases may represent components of a signal transduction pathway regulating IB-␣ levels in vascular endothelium.

Induction of Tissue Factor Expression in Human Endothelial Cells by CD40 Ligand Is Mediated via Activator Protein 1, Nuclear Factor κB, and Egr-1

“…Nevertheless, CD40L-induced phosphorylation of bound c-Jun by the stressactivated protein kinase c-Jun NH 2 -terminal kinase (26) might contribute to increased transcriptional activity of c-Jun (27). The CD40L-dependent activation of NF-B binding to the TF promoter most likely depends on the inactivation of IB␣ via IKK as demonstrated for other cytokines in various cell types, including EC (28,29). The requirement for members of both the AP-1 and NF-B/Rel transcription factor family to maximally induce the TF promoter by CD40L might indicate the necessity of cooperative binding of c-Jun/JunD to the AP-1 sites and c-Rel/p65 to the NF-B site, both located closely in the LRE of the TF promoter.…”

Novel Inhibitors of Cytokine-induced IκBα Phosphorylation and Endothelial Cell Adhesion Molecule Expression Show Anti-inflammatory Effects in Vivo

“…The activities of these kinases were unaffected by 15 min treatment with TNF␣ or by compound 1 (20 M). The molecular weights of these kinases correspond to those expected for the catalytic subunits of casein kinase II; however, Bennett et al (47) have recently described IB-␣ kinases of similar molecular weight in endothelial cells that are distinct from casein kinase II. Our results suggest that compound 1 had no effect on the activity of these IB-␣ kinases.…”

Interleukins in Atherosclerosis: Molecular Pathways and Therapeutic Potential

“…The endothelial NF-B/IB system is presumed to be primed in endothelial cells in lesion-prone arterial sites, as evidenced by increased expression of the p65 (RelA) NF-B subunit, IB␣, and IB␤, prior to plaque development and NF-B activation (Hajra et al, 2000). The attenuation of IB activity by IKK up-regulation has been identified as a pivotal step in endothelial activation (Read et al, 1994;Bennett et al, 1996;Johnson et al, 1996), whereas the inhibition of endothelial adhesion molecule expression by nitric oxide has been found to be mediated by IB␣ (Spiecker et al, 1997). The recognition of the physiological importance of this inhibitory pathway has prompted the evaluation of the anti-atherogenic properties of IB␣ administration.…”