title: Activation of NF-kappaB nuclear transcription factor by flow in human endothelial cells.
abstract: The tractive force generated by blood flow, called fluid shear stress, is an important regulator of endothelial cell gene expression. Several transcription factors are activated by shear stress, including members of the NF-kappaB/Rel family. The nature of the upstream-signaling components involved in the activation of NF-kappaB by flow has been studied in human endothelial cells. Flow rapidly increased endogenous IKK1/2 activity and transiently degraded IkappaBalpha and IkappaBbeta1, but not p105/p50. Nuclear translocation of the p65 subunit was induced by flow in wild-type (w/t) cells and in cells overexpressing w/t NIK, IKK1 or IKK2, but not in cells transiently transfected with kinase-inactive mutants of these enzymes. Nuclear translocation of p65 in response to flow was not affected by overexpressing a dominant-negative mutant of a MAPKKK related to NIK, called TPL2 kinase, nor by pretreating cells with the selective PKC inhibitor bisindoylmaleimide-1. Gel shift assays showed that the binding of p50/p65 heterodimer to radiolabeled oligonucleotide containing a shear-stress response element was increased by flow. The activity of a 3kappaB conA-luciferase reporter was also increased, confirming that NF-kappaB activated by flow was transcriptionally active. We conclude that shear stress induces gene transactivation by NF-kappaB (p50/p65) via the NIK-IKK1/2 pathway and proteosome-dependent degradation of IkappaB and that induction by flow does not involve TPL-2 kinase or PKC.
authors.affiliation: Centre for Biomolecular Sciences, University of St. Andrews, St. Andrews, Fife KY16 9TS, UK.
authors.forename: David C
authors.initials: DC
authors.lastname: Hay
authors.forename: Catherine
authors.initials: C
authors.lastname: Beers
authors.forename: Vicky
authors.initials: V
authors.lastname: Cameron
authors.forename: Lesley
authors.initials: L
authors.lastname: Thomson
authors.forename: Frederick W
authors.initials: FW
authors.lastname: Flitney
authors.forename: Ronald T
authors.initials: RT
authors.lastname: Hay
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000502:physiology; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000235:genetics*; D006439:Hemodynamics / Q000502:physiology; D006801:Humans; D051550:I-kappa B Kinase; D020672:I-kappa B Proteins / Q000378:metabolism; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D012039:Regional Blood Flow / Q000502:physiology; D033081:Shear Strength; D013314:Stress, Mechanical; D015533:Transcriptional Activation / Q000502:physiology*; D019892:eIF-2 Kinase / Q000378:metabolism
chemical_list: D020672:I-kappa B Proteins; D016328:NF-kappa B; D017346:Protein Serine-Threonine Kinases; D019892:eIF-2 Kinase; C496559:CHUK protein, human; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; C496565:IKBKE protein, human; D011493:Protein Kinase C; D020930:MAP Kinase Kinase Kinases