Abstract
Over recent decades, diverse intracellular organelles have been recognized as key determinants of Ca(2+) signaling in eukaryotes. In yeast however, information on intra-organellar Ca(2+) concentrations is scarce, despite the demonstrated importance of Ca(2+) signals for this microorganism. Here, we directly monitored free Ca(2+) in the lumen of the endoplasmic reticulum (ER) of yeast cells, using a specifically targeted version of the Ca(2+)-sensitive photoprotein aequorin. Ca(2+) uptake into the yeast ER displayed characteristics distinctly different from the mammalian ER. At steady-state, the free Ca(2+) concentration in the ER lumen was limited to approximately 10 microM, and ER Ca(2+) sequestration was insensitive to thapsigargin, an inhibitor specific for mammalian ER Ca(2+) pumps. In pmr1 null mutants, free Ca(2+) in the ER was reduced by 50%. Our findings identify the secretory pathway pump Pmr1, predominantly localized in the Golgi, as a major component of ER Ca(2+) uptake activity in yeast.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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ATP-Binding Cassette Transporters / metabolism*
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Aequorin / metabolism
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Calcimycin / pharmacology
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Calcium / metabolism*
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Calcium / pharmacokinetics
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Calcium-Transporting ATPases*
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Cycloheximide / pharmacology
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Endoplasmic Reticulum / metabolism*
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Enzyme Inhibitors / pharmacology
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Golgi Apparatus / metabolism
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Homeostasis
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Ionophores / pharmacology
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Models, Biological
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Molecular Chaperones
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Protein Synthesis Inhibitors / pharmacology
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Recombinant Fusion Proteins / metabolism
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Saccharomyces cerevisiae / metabolism
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Saccharomyces cerevisiae Proteins*
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Sarcoplasmic Reticulum / metabolism*
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Temperature
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Thapsigargin / pharmacology
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Vanadates / pharmacology
Substances
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ATP-Binding Cassette Transporters
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Enzyme Inhibitors
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Ionophores
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Molecular Chaperones
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Protein Synthesis Inhibitors
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Recombinant Fusion Proteins
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SSC1 protein, S cerevisiae
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Saccharomyces cerevisiae Proteins
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Calcimycin
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Vanadates
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Aequorin
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Thapsigargin
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Cycloheximide
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Calcium-Transporting ATPases
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Calcium