Effects of PMCA and SERCA pump overexpression on the kinetics of cell Ca(2+) signalling.
The dynamic interactions of the main pathways for active Ca(2+) transport have been analysed in living cells by altering the expression of their components. The plasma membrane (PMCA) and the endoplasmic reticulum (ER) (SERCA) Ca(2+) pumps were transiently overexpressed in CHO cells, and the Ca(2+) homeostasis in the subcellular compartments was investigated using specifically targeted chimaeras of the Ca(2+)- sensitive photoprotein aequorin. In resting cells, overexpression of the PMCA and SERCA pumps caused a reduction and an increase in ER [Ca(2+)] levels, respectively, while no significant differences were detected in cytosolic and mitochondrial [Ca(2+)]. Upon stimulation with an inositol 1,4, 5-trisphosphate (IP(3))-generating agonist, the amplitude of the mitochondrial and cytosolic Ca(2+) rises correlated with the ER [Ca(2+)] only up to a threshold value, above which the feedback inhibition of the IP(3) channel by Ca(2+) appeared to be limiting.
Brini
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Department of Biochemistry and Center for the Study of Biomembranes of the National Research Council (CNR), University of Padova, Viale G. Colombo, 3, 35121 Padova, Italy. brini@civ.bio.unipd.it
Bano
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Manni
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Rizzuto
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Carafoli
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D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000331:Aequorin / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism*; D002122:Calcium Chloride / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D027682:Cation Transport Proteins; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D003600:Cytosol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005456:Fluorescent Dyes / Q000494:pharmacology; D016257:Fura-2 / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D007700:Kinetics; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism; D053499:Plasma Membrane Calcium-Transporting ATPases; D010957:Plasmids / Q000378:metabolism; D020033:Protein Isoforms; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D015398:Signal Transduction; D014162:Transfection
D027682:Cation Transport Proteins; D005456:Fluorescent Dyes; D020033:Protein Isoforms; D000331:Aequorin; D015544:Inositol 1,4,5-Trisphosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D053499:Plasma Membrane Calcium-Transporting ATPases; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D002122:Calcium Chloride; D002118:Calcium; D016257:Fura-2