Mitochondrial and ER-targeted eCALWY probes reveal high levels of free Zn2+.
ACS Chem Biol
; 9(9): 2111-20, 2014 Sep 19.
Article
em En
| MEDLINE
| ID: mdl-25011072
ABSTRACT
Zinc (Zn2+) ions are increasingly recognized as playing an important role in cellular physiology. Whereas the free Zn2+ concentration in the cytosol has been established to be 0.1-1 nM, the free Zn2+ concentration in subcellular organelles is not well-established. Here, we extend the eCALWY family of genetically encoded Förster Resonance Energy Transfer (FRET) Zn2+ probes to permit measurements in the endo(sarco)plasmic reticulum (ER) and mitochondrial matrix. Deployed in a variety of mammalian cell types, these probes reveal resting mitochondrial free [Zn2+] values of â¼300 pM, somewhat lower than in the cytosol but 3 orders of magnitude higher than recently reported using an alternative FRET-based sensor. By contrast, free ER [Zn2+] was found to be ≥5 nM, which is >5000-fold higher than recently reported but consistent with the proposed role of the ER as a mobilizable Zn2+ store. Treatment of ß-cells or cardiomyocytes with sarco(endo)plasmic reticulum Ca2+-ATPase inhibitors, mobilization of ER Ca2+ after purinergic stimulation with ATP, or manipulation of ER redox, exerted no detectable effects on [Zn2+]ER. These findings question the previously proposed role of Ca2+ in Zn2+ mobilization from the ER and suggest that high ER Zn2+ levels may be an important aspect of cellular homeostasis.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Zinco
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Transferência Ressonante de Energia de Fluorescência
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Retículo Endoplasmático
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Mitocôndrias
Limite:
Animals
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Humans
Idioma:
En
Ano de publicação:
2014
Tipo de documento:
Article