title: Calcineurin/NFAT signaling represses genes Vamp1 and Vamp2 via PMCA-dependent mechanism during dopamine secretion by Pheochromocytoma cells.
abstract: BACKGROUND
  Plasma membrane Ca(2+)-ATPases (PMCA) extrude Ca(2+) 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 [Ca(2+)]c in PC12 cells is maintained mainly by two Ca(2+)-ATPases, PMCA2 and PMCA3. Recently, we found that Ca(2+) 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 Ca(2+)/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.
  
  
  RESULTS
  PMCA2- and PMCA3-deficient cells displayed profound decrease in dopamine secretion accompanied by a permanent increase in [Ca(2+)]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).
  
  
  CONCLUSIONS
  PMCA2 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.
authors.affiliation: Department of Biochemistry, Nencki Institute of Experimental Biology, PAS, Warsaw, Poland; Laboratory of Neurogenetics, Department of Neurodegenerative Disorders, Mossakowski Medical Research Centre PAS, Warsaw, Poland.
authors.forename: Michalina
authors.initials: M
authors.lastname: Kosiorek
authors.affiliation: Department of Molecular Neurochemistry, Medical University, Lodz, Poland.
authors.forename: Ludmila
authors.initials: L
authors.lastname: Zylinska
authors.affiliation: Department of Biochemistry, Nencki Institute of Experimental Biology, PAS, Warsaw, Poland.
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Zablocki
authors.affiliation: Department of Biochemistry, Nencki Institute of Experimental Biology, PAS, Warsaw, Poland.
authors.forename: Slawomir
authors.initials: S
authors.lastname: Pikula
mesh_terms: D000310:Adrenal Gland Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D000818:Animals; D015153:Blotting, Western; D019703:Calcineurin / Q000235:genetics / Q000378:metabolism*; D020013:Calcium Signaling; D002450:Cell Communication; D047369:Chromatin Immunoprecipitation; D004298:Dopamine / Q000378:metabolism*; D005786:Gene Expression Regulation; D008156:Luciferases / Q000378:metabolism; D050778:NFATC Transcription Factors / Q000235:genetics / Q000378:metabolism*; D016716:PC12 Cells; D010673:Pheochromocytoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D053499:Plasma Membrane Calcium-Transporting ATPases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D050987:Vesicle-Associated Membrane Protein 1 / Q000235:genetics / Q000378:metabolism*; D050988:Vesicle-Associated Membrane Protein 2 / Q000235:genetics / Q000378:metabolism*
chemical_list: C000598609:Atp2b3 protein, rat; D050778:NFATC Transcription Factors; D012333:RNA, Messenger; C494706:Vamp2 protein, rat; D050987:Vesicle-Associated Membrane Protein 1; D050988:Vesicle-Associated Membrane Protein 2; C494705:vesicle-associated membrane protein 1, rat; D008156:Luciferases; D019703:Calcineurin; D053499:Plasma Membrane Calcium-Transporting ATPases; D004298:Dopamine