Regulation of the nuclear export of the transcription factor NFATc1 by protein kinases after slow fibre type electrical stimulation of adult mouse skeletal muscle fibres

The transcription factor nuclear factor of activated T cells (NFAT)c1 has been shown to be involved in turning on slow skeletal muscle fibre gene expression. Previous studies from our laboratory have characterized the stimulation pattern-dependent nuclear import and resting shuttling of NFATc1-green fluorescent protein (GFP) in flexor digitorum brevis (FDB) muscle fibres from adult mouse. In this study, we use viral expression of the transcription factor NFATc1-GFP fusion protein to investigate the mechanisms underlying the nuclear export of the NFATc1-GFP that accumulated in the nuclei of cultured dissociated adult mouse FDB muscle fibres during slow-twitch fibre type electrical stimulation. In these studies, we found that inhibition of either glycogen synthase kinase 3 beta (GSK3 beta) or casein kinase 1 or 2 (CK1/2) markedly slowed the decay of nuclear NFATc1-GFP after cessation of muscle fibre electrical stimulation, whereas inhibition of casein kinase 1 delta, p38 mitogen-activated protein kinase, c-Jun N-terminal kinase and protein kinase A had little effect. Simultaneous inhibition of GSK3 beta and CK1/2 completely blocked the nuclear export of NFATc1-GFP after muscle activity. We also developed a simplified model of NFATc1 phosphorylation/dephosphorylation and nuclear fluxes, and used this model to simulate the observed time courses of nuclear NFATc1-GFP with and without NFATc1 kinase inhibition. Our results suggest that GSK3 beta and CK1/2 are the major protein kinases that contribute to the removal of NFATc1 that accumulates in muscle fibre nuclei during muscle activity, and that GSK3 beta and CK1/2 are responsible for phosphorylating NFATc1 in muscle nuclei in a complementary or synergistic fashion.