Store-operated Ca2+ influx and subplasmalemmal mitochondria. Cell Calcium

Section on Molecular Signal Transduction, Program on Developmental Neuroscience, NICHD, NIH, Bethesda, MD, USA.
Cell calcium (Impact Factor: 3.51). 06/2009; 46(1):49-55. DOI: 10.1016/j.ceca.2009.04.002
Source: PubMed


Calcium depletion of the endoplasmic reticulum (ER) induces oligomerisation, puncta formation and translocation of the ER Ca(2+) sensor proteins, STIM1 and -2 into plasma membrane (PM)-adjacent regions of the ER, where they activate the Orai1, -2 or -3 proteins present in the opposing PM. These proteins form ion channels through which store-operated Ca(2+) influx (SOC) occurs. Calcium ions exert negative feed-back on SOC. Here we examined whether subplasmalemmal mitochondria, which reduce this feed-back by Ca(2+) uptake, are located within or out of the high-Ca(2+) microdomains (HCMDs) formed between the ER and plasmalemmal Orai1 channels. For this purpose, COS-7 cells were cotransfected with Orai1, STIM1 labelled with YFP or mRFP and the mitochondrially targeted Ca(2+) sensitive fluorescent protein inverse-Pericam. Depletion of ER Ca(2+) with ATP+thapsigargin (in Ca(2+)-free medium) induced the appearance of STIM1 puncta in the < or =100 nm wide subplasmalemmal space, as examined with TIRF. Mitochondria were located either in the gaps between STIM1-tagged puncta or in remote, STIM1-free regions. After addition of Ca(2+) mitochondrial Ca(2+) concentration increased irrespective of the mitochondrion-STIM1 distance. These observations indicate that mitochondria are exposed to Ca(2+) diffused laterally from the HCMDs formed between the PM and the subplasmalemmal ER.

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Available from: Marek Korzeniowski, Jan 12, 2014
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    • "Interestingly, a considerable fraction of the peri-PM mitochondria also appear to be coupled to the PM through an ER stack. Of note, this indicates the association of mitochondria with the ER subdomains that control store-operated Ca 2+ entry (Korzeniowski et al., 2009; Wu et al., 2006) and provides the structural basis for the observation that the entering Ca 2+ crosses the ER before it is taken up by the mitochondria (Demaurex et al., 2009). At the level of individual mitochondria, the contacts are heterogeneous and commonly appear as discrete spots of various size and shape. "
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    ABSTRACT: The ER-mitochondrial junction provides a local calcium signaling domain that is critical for both matching energy production with demand and the control of apoptosis. Here, we visualize ER-mitochondrial contact sites and monitor the localized [Ca(2+)] changes ([Ca(2+)](ER-mt)) using drug-inducible fluorescent interorganelle linkers. We show that all mitochondria have contacts with the ER, but plasma membrane (PM)-mitochondrial contacts are less frequent because of interleaving ER stacks in both RBL-2H3 and H9c2 cells. Single mitochondria display discrete patches of ER contacts and show heterogeneity in the ER-mitochondrial Ca(2+) transfer. Pericam-tagged linkers revealed IP(3)-induced [Ca(2+)](ER-mt) signals that exceeded 9 microM and endured buffering bulk cytoplasmic [Ca(2+)] increases. Altering linker length to modify the space available for the Ca(2+) transfer machinery had a biphasic effect on [Ca(2+)](ER-mt) signals. These studies provide direct evidence for the existence of high-Ca(2+) microdomains between the ER and mitochondria and suggest an optimal gap width for efficient Ca(2+) transfer.
    Molecular cell 07/2010; 39(1):121-32. DOI:10.1016/j.molcel.2010.06.029 · 14.02 Impact Factor
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    • "This conclusion is also consistent with recent morphological data (Korzeniowski et al., 2009). It is suggested that previous models in which mitochondria were thought to play a local Ca 2+ buffering role at the mouth of CCE channels must be revisited (Hoth et al., 1997; Parekh, 2003). "
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    ABSTRACT: Although it is widely accepted that mitochondria in living cells can efficiently uptake Ca(2+) during stimulation because of their vicinity to microdomains of high [Ca(2+)], the direct proof of Ca(2+) hot spots' existence is still lacking. Thanks to a GFP-based Ca(2+) probe localized on the cytosolic surface of the outer mitochondrial membrane, we demonstrate that, upon Ca(2+) mobilization, the [Ca(2+)] in small regions of the mitochondrial surface reaches levels 5- to 10-fold higher than in the bulk cytosol. We also show that the [Ca(2+)] to which mitochondria are exposed during capacitative Ca(2+) influx is similar between near plasma membrane mitochondria and organelles deeply located in the cytoplasm, whereas it is 2- to 3-fold higher in subplasma membrane mitochondria upon activation of voltage-gated Ca(2+) channels. These results demonstrate that mitochondria are exposed to Ca(2+) hot spots close to the ER but are excluded from the regions where capacitative Ca(2+) influx occurs.
    Molecular cell 04/2010; 38(2):280-90. DOI:10.1016/j.molcel.2010.04.003 · 14.02 Impact Factor
  • Journal of the American College of Cardiology 02/1995; 25(2). DOI:10.1016/0735-1097(95)93150-B · 16.50 Impact Factor
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