Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Deak, A.
New aspects of STIM1 signaling: Stimulus-specific contribution of mitochondrial Ca2+ uptake to STIM1 activation and SOCE
PhD-Studium (Doctor of Philosophy); Humanmedizin; [ Dissertation ] Medical University of Graz; 2014. pp. 103
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- Autor*innen der Med Uni Graz:
- Deak Andras Tamas
- Betreuer*innen:
- Graier Wolfgang
- Malli Roland
- Altmetrics:
- Abstract:
- Store-operated Ca2+ entry (SOCE) is a ubiquitous Ca2+ influx pathway in mammalian cells activated following depletion of the endoplasmic reticulum (ER) Ca2+ stores. The essential physiological role of SOCE is the replenishment of depleted Ca2+ pools and the generation of sustained cytosolic Ca2+ signals required for fundamental cellular functions such as the secretion of vesicle contents, cytoskeleton remodeling or gene transcription. It is now recognized that deficiencies in SOCE may also associated with pathomechanisms of certain diseases including immunodeficiency, cardiac hypertrophy or Alzheimer´s disease. The molecular mechanism of SOCE activation is virtually based on the complex interaction of the ER-spanning stromal interacting molecule (STIM) 1 protein and plasma membrane (PM) Ca2+ channels such as Orai or transient receptor potential cation channels (TRPC). STIM1 serves as the central multifunctional “relay” in the activation mechanism of SOCE. It acts as a “detector” by sensing ER Ca2+ content through its luminal Ca2+ binding domain. STIM1 functions also as a “courier” by migrating across the ER to subplasmalemmal areas and delivers the message to the PM that the Ca2+ stores are empty. Moreover, STIM1 is an “operator” of SOCE, as it directly binds to and activates store-operated Ca2+ channels. Although the activation mechanism and the components of the SOCE machinery are well described, the regulation of SOCE is still under extensive research. One part of this research focuses on the understanding how mitochondria control SOCE. Even though, it has been established long ago that mitochondrial Ca2+ uptake contributes to SOCE regulation, the exact mechanisms, how this is accomplished, are still insufficiently clarified. The unclear results partly originate from the fact that proteins mediating mitochondrial Ca2+ uptake [i.e. mitochondrial Ca2+ uniporter (MCU), mitochondrial calcium uptake 1 (MICU1), uncoupling protein 2/3 (UCP2/3), leucine zipper-EF-hand containing transmembrane protein 1 (Letm1)] have been identified only recently. Thus, the contribution of mitochondrial Ca2+ uptake to SOCE has been investigated indirectly until now. This work was designed to elucidate the molecular mechanisms underlying the mitochondrial control of SOCE. For this purpose a new technique was established, which enabled the simultaneous recording of mitochondrial and cytosolic Ca2+ signals in the same individual cell with fluorescent microscopy. With a series of organelle-associated Ca2+ measurements using a newly constructed mitochondrial Ca2+ probe (4mtD1GO), evidence was found for distinct modes of mitochondrial Ca2+ uptake depending on sarco/endoplasmic reticulum Ca2+ ATPase (SERCA) activity. Taking advantage of these findings, the direct involvement of mitochondrial Ca2+ buffering in SOCE was also investigated. These experiments were performed in HeLa cells stably depleted of either MCU or UCP2. The lack of either of these two proteins results in decelerated STIM1 activation and impaired SOCE following cell stimulation with an IP3-generating agonist. Upon artificially augmented cytosolic Ca2+-buffering or ER Ca2+ depletion by SERCA inhibitors, STIM1 activation did not rely on intact mitochondrial Ca2+ uptake. However, MCU-dependent mitochondrial sequestration of Ca2+ entering through the SOCE pathway was essential to prevent slow deactivation of SOCE. In summary, the findings herein support the existence of distinct mitochondrial Ca2+ uptake routes and highlight a special and tight regulation of STIM1 activation and SOCE by mitochondria. Considering the central role of STIM1, the identification of any molecular mechanisms that regulate this protein under physiological conditions of cell stimulation will help to improve our understanding of other STIM1-dependent cell signaling events as well.