Abstract
BackgroundPlasma membrane Ca2+-ATPases (PMCA) extrude Ca2+ ions out of the cell and contribute to generation of calcium oscillations. Calcium signaling is crucial for transcriptional regulation of dopamine secretion by neuroendocrine PC12 cells. Low resting [Ca2+]c in PC12 cells is maintained mainly by two Ca2+-ATPases, PMCA2 and PMCA3. Recently, we found that Ca2+ dependent phosphatase calcineurin was excessively activated under conditions of experimental downregulation of PMCA2 or PMCA3. Thus, the aim of this study was to explain if, via modulation of the Ca2+/calcineurin-dependent nuclear factor of activated T cells (NFAT) pathway, PMCA2 and PMCA3 affect intracellular signaling in pheochromocytoma/neuronal cells/PC12 cells. Secondly, we tested whether this might influence dopamine secretion by PC12 cells.ResultsPMCA2- and PMCA3-deficient cells displayed profound decrease in dopamine secretion accompanied by a permanent increase in [Ca2+]c. Reduction in secretion might result from changes in NFAT signaling, following altered PMCA pattern. Consequently, activation of NFAT1 and NFAT3 transcription factors was observed in PMCA2- or PMCA3-deficient cells. Furthermore, chromatin immunoprecipitation assay indicated that NFATs could be involved in repression of Vamp genes encoding vesicle associated membrane proteins (VAMP).ConclusionsPMCA2 and PMCA3 are crucial for dopamine secretion in PC12 cells. Reduction in PMCA2 or PMCA3 led to calcium-dependent activation of calcineurin/NFAT signaling and, in consequence, to repression of the Vamp gene and deterioration of the SNARE complex formation in PC12 cells.
Highlights
Pheochromocytoma is a tumor characterized by an excessive catecholamine secretion [1]
The quantitative PCR (qPCR) results are in agreement with the protein level, showing that in PMCA3-deficient cells there was a slight increase in a protein content of the PMCA2 isoform
As we have already reported [15], downregulation of PMCA2 or PMCA3 isoforms resulted in a permanent elevation of [Ca2+]c under resting conditions (5 mM KCl) and an increase in calcium influx upon plasma membrane depolarization in the presence of 59 mM KCl supplier
Summary
Pheochromocytoma is a tumor characterized by an excessive catecholamine secretion [1]. One of the catecholamines excessively secreted during progression of this tumor, is dopamine. This is a neurotransmitter and neurohormone known to be involved in a variety of processes in the brain, including cognition, learning, attention, reward system, control of emotions and motor coordination [2]. In this study, using PC12 cells as a model, we focused on a2+-dependent signaling during dopamine secretion in dopaminergic tumor pheochromocytoma. We found that Ca2+ dependent phosphatase calcineurin was excessively activated under conditions of experimental downregulation of PMCA2 or PMCA3. The aim of this study was to explain if, via modulation of the Ca2+/calcineurindependent nuclear factor of activated T cells (NFAT) pathway, PMCA2 and PMCA3 affect intracellular signaling in pheochromocytoma/neuronal cells/PC12 cells. We tested whether this might influence dopamine secretion by PC12 cells
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