The transcription factors NFAT and CREB have different susceptibilities to the reduced ca responses caused by the knock down of inositol trisphosphate receptor in HEK 293A cells

Abstract

Background/Aims: The inositol 1,4,5-trisphosphate receptor (IP 3 R), a ligand-gated Ca 2+ channel, plays an important role in the control of intracellular Ca 2+ . Three isoforms of IP 3 R have been identified and most cell types express different proportions of these isoforms. The purpose of this study was to investigate how IP 3 R signalling is involved in the activation of the Ca 2+ -sensitive transcription factors NFAT and CREB. Methods: Each IP 3 R isoform expressed in HEK 293A cells was knocked down using selective siRNA. Free intracellular Ca 2+ was monitored spectrofluometrically. NFAT and CREB activities were measured with luciferase reporter constructs. Results: IP 3 R-2-knocked down HEK 293A cells showed a deficient CCh-induced Ca 2+ 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 3 R-2-knocked down HEK 293A cells. Overexpression of IP 3 R-1 could fully compensate for IP 3 R-2 knock down to mobilize Ca 2+ and to activate NFAT. Conclusion: Our results show that the knock down of IP 3 R-2 significantly reduced the intracellular Ca 2+ response of HEK 293 cells. This reduced Ca 2+ response did not affect the activation of CREB but significantly decreased the activation of NFAT, suggesting that the Ca 2+ signals required for the activation of NFAT are stronger than those required for the activation of CREB. © 2010 S. Karger AG Basel.