NFAT activation by membrane potential follows a calcium pathway distinct from other activity-related transcription factors in skeletal muscle cells

Depolarization of skeletal muscle cells triggers intracellular calcium signals mediated by ryanodine and IP3 receptors. We have reported that K+-induced depolarization activates the transcriptional regulators ERKs, CREB, c-fos, c-jun and egr-1 through IP3-dependent calcium release, whereas NF B activation is elicited by both ryanodine and IP3 receptors-mediated calcium signals. We have further showed that field stimulation with electrical pulses results in an NF B activation increase being it dependent of the amount of pulses and independent of their frequency. In this work, we report the results obtained for NFAT-mediated transcription and translocation generated by both K+ and electrical stimulation protocols in primary skeletal muscle cells and in C2C12 cells. The calcium source for NFAT activation is through release by ryanodine receptors and extracellular calcium entry. We found this activation to be independent on the number of pulses within a physiological range of stimulus frequency and enhanced by long-lasting low frequency stimulation. Therefore, activation of NFAT signaling pathway differs from that of NF B and other transcription factors. Calcineurin enzyme activity correlates well with the relative activation of NFAT translocation and transcription using different stimulation protocols. Furthermore, both K+-induced depolarization and electrical stimulation increases mRNA levels of type 1 IP3 receptor mediated by calcineurin activity, which suggests that depolarization may regulate IP3 receptor transcription. These results confirm the presence of at least two independent pathways for excitationtranscription coupling in skeletal muscle cells, both dependent on calcium release and triggered by the same voltage sensor but activating different intracellular release channels.This work was supported by Fondo Nacional de Investigación Científica y Tecnológica (FONDECYT) grants numbers 1030988 and 1060177, and by Fondo Nacional de Investigación en Areas Prioritarias (FONDAP) Center for Molecular Studies of the Cell, grant number 15010006