Ca2+ entry via TRPC channels is necessary for thrombin-induced NF-kappaB activation in endothelial cells through AMP-activated protein kinase and protein kinase Cdelta

• Published in: The Journal of biological chemistry 2009-01, Vol.284 (1), p.563-574
• Main Authors: Bair, Angela M, Thippegowda, Prabhakar B, Freichel, Marc, Cheng, Ni, Ye, Richard D, Vogel, Stephen M, Yu, Yanni, Flockerzi, Veit, Malik, Asrar B, Tiruppathi, Chinnaswamy
• Format: Article
• Language: English
• Subjects: AMP-Activated Protein Kinases - genetics; AMP-Activated Protein Kinases - metabolism; Animals; Calcium; Calcium-Calmodulin-Dependent Protein Kinases - genetics; Calcium-Calmodulin-Dependent Protein Kinases - metabolism; Cells, Cultured; Endothelial Cells - cytology; Endothelial Cells - metabolism; Enzyme Activation - drug effects; Hemostatics - pharmacology; Humans; Mice; Mice, Knockout; Protein Kinase C-delta - genetics; Protein Kinase C-delta - metabolism; Signal Transduction - drug effects; Signal Transduction - genetics; Thrombin - pharmacology; Thrombosis - genetics; Thrombosis - metabolism; Transcription Factor RelA - genetics; Transcription Factor RelA - metabolism; TRPC Cation Channels - genetics; TRPC Cation Channels - metabolism
• ISSN: 0021-9258
• DOI: 10.1074/jbc.M803984200

The transient receptor potential canonical (TRPC) family channels are proposed to be essential for store-operated Ca2+ entry in endothelial cells. Ca2+ signaling is involved in NF-kappaB activation, but the role of store-operated Ca2+ entry is unclear. Here we show that thrombin-induced Ca2+ entry and the resultant AMP-activated protein kinase (AMPK) activation targets the Ca2+-independent protein kinase Cdelta (PKCdelta) to mediate NF-kappaB activation in endothelial cells. We observed that thrombin-induced p65/RelA, AMPK, and PKCdelta activation were markedly reduced by knockdown of the TRPC isoform TRPC1 expressed in human endothelial cells and in endothelial cells obtained from Trpc4 knock-out mice. Inhibition of Ca2+/calmodulin-dependent protein kinase kinase beta downstream of the Ca2+ influx or knockdown of the downstream Ca2+/calmodulin-dependent protein kinase kinase beta target kinase, AMPK, also prevented NF-kappaB activation. Further, we observed that AMPK interacted with PKCdelta and phosphorylated Thr505 in the activation loop of PKCdelta in thrombin-stimulated endothelial cells. Expression of a PKCdelta-T505A mutant suppressed the thrombin-induced but not the TNF-alpha-induced NF-kappaB activation. These findings demonstrate a novel mechanism for TRPC channels to mediate NF-kappaB activation in endothelial cells that involves the convergence of the TRPC-regulated signaling at AMPK and PKCdelta and that may be a target of interference of the inappropriate activation of NF-kappaB associated with thrombosis.