Na+/Ca2+ exchanger overexpression impairs calcium signaling in fibroblasts: inhibition of the [Ca2+] increase at the cell periphery and retardation of cell adhesion.
We examined the Ca2+ handling property and cell function of CCL39 fibroblasts highly overexpressing the cardiac isoform (NCX1) of Na+/ Ca2+ exchanger. In NCX1 transfectants in 146 mM Na+, ionomycin, alpha-thrombin or thapsigargin only produced a small transient increase in [Ca2+]i compared to the large increase seen in control cells, although resting [Ca2+]i was not significantly different between these cells. In Na+-free medium, in contrast, the [Ca2+]i responses in NCX1 transfectants and control cells stimulated with these agents were not different, indicating that the Ca2+ content of the intracellular store(s) does not decrease on NCX1 transfection. The expression levels of the endoplasmic reticulum and plasma membrane Ca2+-ATPases, and thrombin- or serum-stimulated cell growth were not altered in NCX1 transfectants. The latter finding suggests that Ca2+ signaling in the nucleus is not impaired appreciably. On fluorescence imaging and confocal microscopy, we found that [Ca2+] did not increase in the peripheral cytoplasm of these cells treated with alpha-thrombin in Na+-containing medium. In these NCX1 transfectants, activation of the plasma membrane Ca2+-activated K+ channels by thrombin or ionomycin was markedly suppressed, and the integrin-mediated adhesion to substrate was significantly delayed compared with control cells. NCX1-overexpressing CCL39 cells thus seem to be a good model with which we can study the Ca2+-regulated membrane processes under physiologically relevant conditions.
Department of Molecular Physiology, National Cardiovascular Center Research Institute, Suita, Osaka/Japan.
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Iwamoto
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Wakabayashi
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Imagawa
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Shigekawa
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D002448:Cell Adhesion; D002455:Cell Division; D002460:Cell Line; D005347:Fibroblasts / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D019831:Sodium-Calcium Exchanger / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
D011994:Recombinant Proteins; D019831:Sodium-Calcium Exchanger; C110374:sodium-calcium exchanger 1; D002118:Calcium