NFAT and NF-kB are important transcriptional regulators that play a role in controlling the expression of several cytokines, cytokine receptors and adhesion molecules. The effect of extracellular ATP on the activation of NF-kB and NFAT was investigated in the mouse microglial cell lines N9 and N13. ATP potently activated NF-kB and NFAT with strikingly distinct kinetics. NFAT was induced as soon as 1 min after stimulation with the nucleotide, while NF-kB activation was quite delayed (3 h). Both NFAT-1 and NFAT-2 were activated by a calcineurin-dependent pathway that required influx of extracellular Ca2+. Inhibition of calcineurin and Ca2+ chelation prevented ATP-induced NFAT activation and nuclear translocation. Exposure to LPS induced the prototypical NF-kB p50 homo and p65/p50 heterodimers while ATP, only induced the peculiar p65 homodimer. NFAT and NF-kB activation involved a P2X-mediated pathway as oxidized ATP completely inhibited the activation of both factors and BzATP was a better agonist than ATP in inducing NFAT and NF-kB. Cells selected for the absence of P2X7 receptor but still expressing the P2Y subtype were refractory to ATP, suggesting that ATP-stimulated activation was a P2X7-mediated process. Furthermore, NF-kB activation required ROI production and involved proteasome activity, while secretion of soluble proteins was not required. Caspase-1 (ICE) was also activated during exposure to ATP.
Purinoceptor-mediated activation of transcription factors NFAT and NF-kB in microglial cells.
FERRARI, Davide;
2000
Abstract
NFAT and NF-kB are important transcriptional regulators that play a role in controlling the expression of several cytokines, cytokine receptors and adhesion molecules. The effect of extracellular ATP on the activation of NF-kB and NFAT was investigated in the mouse microglial cell lines N9 and N13. ATP potently activated NF-kB and NFAT with strikingly distinct kinetics. NFAT was induced as soon as 1 min after stimulation with the nucleotide, while NF-kB activation was quite delayed (3 h). Both NFAT-1 and NFAT-2 were activated by a calcineurin-dependent pathway that required influx of extracellular Ca2+. Inhibition of calcineurin and Ca2+ chelation prevented ATP-induced NFAT activation and nuclear translocation. Exposure to LPS induced the prototypical NF-kB p50 homo and p65/p50 heterodimers while ATP, only induced the peculiar p65 homodimer. NFAT and NF-kB activation involved a P2X-mediated pathway as oxidized ATP completely inhibited the activation of both factors and BzATP was a better agonist than ATP in inducing NFAT and NF-kB. Cells selected for the absence of P2X7 receptor but still expressing the P2Y subtype were refractory to ATP, suggesting that ATP-stimulated activation was a P2X7-mediated process. Furthermore, NF-kB activation required ROI production and involved proteasome activity, while secretion of soluble proteins was not required. Caspase-1 (ICE) was also activated during exposure to ATP.I documenti in SFERA sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
