The Transcription Factors NFAT and CREB Have Different Susceptibilities to the Reduced Ca2+ Responses Caused by the Knock Down of Inositol Trisphosphate Receptor in HEK 293A Cells
Background/Aims: The inositol 1,4,5-trisphosphate receptor (IP<sub>3</sub>R), a ligand-gated Ca<sup>2+</sup> channel, plays an important role in the control of intracellular Ca<sup>2+</sup>. Three isoforms of IP<sub>3</sub>R have been identified and most cell types express different proportions of these isoforms. The purpose of this study was to investigate how IP<sub>3</sub>R signalling is involved in the activation of the Ca<sup>2+</sup>-sensitive transcription factors NFAT and CREB. Methods: Each IP<sub>3</sub>R isoform expressed in HEK 293A cells was knocked down using selective siRNA. Free intracellular Ca<sup>2+</sup> was monitored spectrofluometrically. NFAT and CREB activities were measured with luciferase reporter constructs. Results: IP<sub>3</sub>R-2-knocked down HEK 293A cells showed a deficient CCh-induced Ca<sup>2+</sup> response that could be rescued by co-stimulation with VIP, a cAMP increasing agonist. NFAT transcriptional activity, but not CREB transcriptional activity, was significantly reduced in IP<sub>3</sub>R-2-knocked down HEK 293A cells. Overexpression of IP<sub>3</sub>R-1 could fully compensate for IP<sub>3</sub>R-2 knock down to mobilize Ca<sup>2+</sup> and to activate NFAT. Conclusion: Our results show that the knock down of IP<sub>3</sub>R-2 significantly reduced the intracellular Ca<sup>2+</sup> response of HEK 293 cells. This reduced Ca<sup>2+</sup> response did not affect the activation of CREB but significantly decreased the activation of NFAT, suggesting that the Ca<sup>2+</sup> signals required for the activation of NFAT are stronger than those required for the activation of CREB.