We have previously demonstrated that Ca²+/calcineurin-dependent dephosphorylation of the transcription factor nuclear factor of activated T cells subtype 1 (NFATc1) during repetitive skeletal muscle activity causes NFAT nuclear translocation and concentration in subnuclear NFAT foci. We now show that NFAT nuclear foci colocalize with heterochromatin regions of intense staining by DAPI or TO-PRO-3 that are present in the nucleus prior to NFATc1 nuclear entry. Nuclear NFATc1 also colocalizes with the heterochromatin markers trimethyl-histone H3 (Lys9) and heterochromatin protein 1α. Mutation of the NFATc1 DNA binding sites prevents entry and localization of NFATc1 in heterochromatin regions. However, fluorescence in situ hybridization shows that the NFAT-regulated genes for slow and fast myosin heavy chains are not localized within the heterochromatin regions. Fluorescence recovery after photobleaching shows that within a given nucleus, NFATc1 redistributes relatively rapidly (t(¹/₂) < 1 min) between NFAT foci. Nuclear export of an NFATc1 mutant not concentrated in NFAT foci is accelerated following nuclear entry during fiber activity, indicating buffering of free nuclear NFATc1 by NFATc1 within the NFAT foci. Taken together, our results suggest that NFAT foci serve as nuclear storage sites for NFATc1, allowing it to rapidly mobilize to other nuclear regions as required.