Steady-state free Ca(2+) in the yeast endoplasmic reticulum reaches only 10 microM and is mainly controlled by the secretory pathway pump pmr1.
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.
Strayle
J
J
Institut für Biochemie der Universität Stuttgart, Pfaffenwaldring 55, D-70569 Stuttgart, Germany.
Pozzan
T
T
Rudolph
H K
HK
D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D000331:Aequorin / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism* / Q000493:pharmacokinetics; D000252:Calcium-Transporting ATPases; D003513:Cycloheximide / Q000494:pharmacology; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006056:Golgi Apparatus / Q000378:metabolism; D006706:Homeostasis; D007476:Ionophores / Q000494:pharmacology; D008954:Models, Biological; D018832:Molecular Chaperones; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D013696:Temperature; D019284:Thapsigargin / Q000494:pharmacology; D014638:Vanadates / Q000494:pharmacology
D018528:ATP-Binding Cassette Transporters; D004791:Enzyme Inhibitors; D007476:Ionophores; D018832:Molecular Chaperones; D011500:Protein Synthesis Inhibitors; D011993:Recombinant Fusion Proteins; C101767:SSC1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D000001:Calcimycin; D014638:Vanadates; D000331:Aequorin; D019284:Thapsigargin; D003513:Cycloheximide; D000252:Calcium-Transporting ATPases; D002118:Calcium