Multiple signaling pathways regulate NF-kappaB-dependent transcription of the monocyte chemoattractant protein-1 gene in primary endothelial cells.
The cytokine-induced C-C chemokine monocyte chemoattractant protein-1 (MCP-1) is an important regulator of leukocyte recruitment to sites of inflammatory challenge. Here, it is demonstrated that the widely distributed contact hapten NiCl(2), like tumor necrosis factor alpha (TNFalpha), induces monocyte-chemoattractant activity in primary human endothelial cells via induction of MCP-1. NiCl(2) rapidly activated mitogen-activated protein (MAP) kinase p38, and inhibition of p38 partially blocked NiCl(2)-induced MCP-1 messenger RNA and protein expression. Both NiCl(2)- and TNFalpha-induced MCP-1 synthesis was sensitive to D609, an inhibitor of phosphatidylcholine-dependent phospholipase C (PC-PLC). NiCl(2)-induced MCP-1 synthesis required activation of NF-kappaB since mutation of NF-kappaB-binding sites in the promoter resulted in complete loss of inducible promoter activity. Consistent with that finding, stimulation with NiCl(2) or TNFalpha activated IkappaB kinase-beta (IKKbeta), and transient transfection of dominant-negative IKKbeta strongly inhibited NiCl(2)- and TNFalpha-induced MCP-1 expression. However, D609 and the specific p38 inhibitor SB202190 did not affect NiCl(2)- and TNFalpha-induced IKKbeta activation, NF-kappaB DNA-binding activity, or transcriptional activity of a Gal4p65 fusion protein. This indicates that p38- and PC-PLC-dependent pathways directly regulate the transcriptional activity of NF-kappaB factors in the transcriptional complex. Consistent with that, inhibition of p38 blocked enhanced transcriptional activity induced by the transcriptional coactivator p300. Thus, it was concluded that at least 3 independent pathways regulate MCP-1 expression in endothelial cells. Its induction requires activation of the IKKbeta/IkappaBalpha/NF-kappaB signaling pathway, resulting in nuclear accumulation of p65 and subsequent recruitment of cofactors. Proper assembly and activity of this transcriptional complex is further modulated by the p38 MAP kinase cascade and a PC-PLC-dependent pathway.
Goebeler
M
M
Klinik und Poliklinik für Hautkrankheiten and the Institut für Medizinische Strahlenkunde und Zellforschung, University of Würzburg, Würzburg, Germany. goebeler-m.derma@mail.uni-wuerzburg.de
Gillitzer
R
R
Kilian
K
K
Utzel
K
K
Bröcker
E B
EB
Rapp
U R
UR
Ludwig
S
S
D018932:Chemokine CCL2 / Q000096:biosynthesis* / Q000235:genetics*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism* / Q000502:physiology; D005786:Gene Expression Regulation / Q000187:drug effects; D006241:Haptens; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology; D016328:NF-kappa B / Q000494:pharmacology*; D009532:Nickel / Q000494:pharmacology; D009687:Nuclear Proteins / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000187:drug effects; D012333:RNA, Messenger / Q000187:drug effects; D015398:Signal Transduction / Q000502:physiology; D015534:Trans-Activators / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D010738:Type C Phospholipases / Q000502:physiology; D014471:Umbilical Veins / Q000166:cytology; D048051:p38 Mitogen-Activated Protein Kinases
D018932:Chemokine CCL2; D006241:Haptens; D016328:NF-kappa B; D009687:Nuclear Proteins; D012333:RNA, Messenger; D015534:Trans-Activators; D014409:Tumor Necrosis Factor-alpha; C022838:nickel chloride; D009532:Nickel; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D010738:Type C Phospholipases