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Gene expression pattern in PC12 cells with reduced PMCA2 or PMCA3 isoform: selective up-regulation of calmodulin and neuromodulin

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Abstract

Cellular calcium homeostasis is controlled predominantly by the plasma membrane calcium pump (PMCA). From four PMCA isoforms, PMCA1 and PMCA4 are ubiquitous, while PMCA2 and PMCA3 are found in excitable cells. We have previously shown that suppression of neuron-specific PMCAs in non-differentiated PC12 cells changed the cell morphology and triggered neuritogenesis. Using the microarrays, real-time PCR and immunodetection, we analyzed the effect of PMCA2 or PMCA3 reduction in PC12 cells on gene expression, with emphasis on calmodulin (CaM), neuromodulin (GAP43) and MAP kinases. In PMCA-suppressed lines total CaM increased, and the calm I and calm II genes appeared to be responsible for this effect. mRNA and protein levels of GAP43 were increased, however, the amount of phosphorylated form was lower than in control cells. Localization of CaM/GAP43 and CaM/pGAP43 differed between control and PMCA-reduced cells. In both PMCA-modified lines, amounts of ERK1/2 increased. While pERK1 decreased, the pERK2 level was similar in all examined lines. PMCA suppression did not change the p38 amount, but the p-p38 diminished. JNK2 protein decreased in both PMCA-reduced cells without changes in pJNK level. Microarray analysis revealed distinct expression patterns of certain genes involved in the regulation of cell cycle, proliferation, migration, differentiation, apoptosis and cell signaling. Suppression of neuron-specific PMCA isoforms affected the phenotype of PC12 cells enabling adaptation to the sustained increase in cytosolic Ca2+ concentration. This is the first report showing function of PMCA2 and PMCA3 isoforms in the regulation of signaling pathways in PC12 cells.

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Abbreviations

CaM:

Calmodulin

CREB:

cAMP Response element-binding

ERK1 and ERK2:

Extracellular signal-regulated kinase 1 and 2

GAP43:

Growth-associated protein 43, neuromodulin

JNK2:

c-Jun N-terminal kinase 2

NFAT:

Nuclear factor of activated T-cells

NGF:

Nerve growth factor

p38 MAPK:

p38 Mitogen-activated protein kinase

PMCA:

Plasma membrane Ca2+-ATPase

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Acknowledgments

We are grateful to Eleonora Welhan for her excellent technical assistance. Supported by the grant 2P05A 03529 from the Ministry of Education and Science, Poland and grants 502-03/6-086/502-64-003 and 503/6-086-02/503-01 from Medical University of Lodz, Poland.

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Authors and Affiliations

  1. Department of Molecular Neurochemistry, Medical University, 6/8 Mazowiecka Street, 92-215, Lodz, Poland

    Tomasz Boczek, Anna Kozaczuk, Bozena Ferenc & Ludmila Zylinska

  2. Department of Biochemistry, Nencki Institute of Experimental Biology, 3 Pasteur Street, 02-093, Warsaw, Poland

    Michalina Kosiorek & Slawomir Pikula

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  1. Tomasz Boczek
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  2. Anna Kozaczuk
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  6. Ludmila Zylinska
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Correspondence to Ludmila Zylinska.

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Boczek, T., Kozaczuk, A., Ferenc, B. et al. Gene expression pattern in PC12 cells with reduced PMCA2 or PMCA3 isoform: selective up-regulation of calmodulin and neuromodulin. Mol Cell Biochem 360, 89–102 (2012). https://doi.org/10.1007/s11010-011-1047-3

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