Functional importance of PMCA isoforms in growth and development of PC12 cells.
Intracellular Ca2+ in neuronal cells is an essential regulatory ion responsible for excitability, synaptic plasticity, and neurite outgrowth. Plasma membrane calcium ATPase (PMCA) is the most sensitive enzyme in decreasing of the Ca2+ concentration. The diverse PMCA isoforms composition in the membranes suggests their specific function in the cell, and whereas PMCA1 and 4 appear to be ubiquitous, PMCA2 and 3 are characteristic isoforms for excitable cells. The aim of our study was to elucidate if and how the elimination of neuron-specific isoforms affects the pattern of cell growth and development. We have obtained stable-transfected PC12 cell lines with a suppressed expression of PMCA2, PMCA3, or both neuron-specific isoforms. The modified profile of PMCA generated considerable changes in morphology of examined PC12 lines, suggesting the activation of a differentiation process to pseudoneuronal phenotype. Experiments with Fura-2/AM-loaded cells revealed an increased cytosolic Ca2+ concentration in the cell lines with blocked PMCA2 isoform. The suppression of PMCA2 concomitantly altered expression of sarco/endoplasmic Ca2+-ATPase 2 isoform (SERCA2) at the protein level. Comparative flow cytometry analysis, using Annexin V/PI conjugate, showed the difference in the mean percentage of apoptotic cells in modified PC12 lines. Our data suggest that specific PMCA isoforms presence can regulate the intact cell development; however, it may involve multiple unidentified yet signaling pathways.
Zylinska
Ludmila
L
Department of Molecular Neurochemistry, Medical University, 6/8 Mazowiecka Street, 92-215 Lodz, Poland. luska@csk.umed.lodz.pl
Kozaczuk
Anna
A
Szemraj
Janusz
J
Kargas
Christos
C
Kowalska
Iwona
I
D000818:Animals; D001483:Base Sequence; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002455:Cell Division; D002462:Cell Membrane / Q000201:enzymology; D017931:DNA Primers; D005434:Flow Cytometry; D007527:Isoenzymes / Q000378:metabolism*; D016716:PC12 Cells; D010957:Plasmids; D051381:Rats
D017931:DNA Primers; D007527:Isoenzymes; D000252:Calcium-Transporting ATPases