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Apoptosis is regulated by the VDAC1 N-terminal region and by VDAC oligomerization: Release of cytochrom c, AIF and Smac/Diablo

Department of Life Sciences, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel.
Biochimica et Biophysica Acta (Impact Factor: 4.66). 03/2010; 1797(6-7):1281-91. DOI: 10.1016/j.bbabio.2010.03.003
Source: PubMed

ABSTRACT Mitochondria, central to basic life functions due to their generation of cellular energy, also serve as the venue for cellular decisions leading to apoptosis. A key protein in mitochondria-mediated apoptosis is the voltage-dependent anion channel (VDAC), which also mediates the exchange of metabolites and energy between the cytosol and the mitochondria. In this study, the functions played by the N-terminal region of VDAC1 and by VDAC1 oligomerization in the release of cytochrome c, Smac/Diablo and apoptosis-inducing factor (AIF) and subsequent apoptosis were addressed. We demonstrate that cells undergoing apoptosis induced by STS or cisplatin and expressing N-terminally truncated VDAC1 do not release cytochrome c, Smac/Diablo or AIF. Ruthenium red (RuR), AzRu, DIDS and hexokinase-I (HK-I), all known to interact with VDAC, inhibited the release of cytochrome c, Smac/Diablo and AIF, while RuR-mediated inhibition was not observed in cells expressing RuR-insensitive E72Q-VDAC1. These findings suggest that VDAC1 is involved in the release of not only cytochrome c but also of Smac/Diablo and AIF. We also demonstrate that apoptosis induction is associated with VDAC oligomerization, as revealed by chemical cross-linking and monitoring in living cells using Bioluminescence Resonance Energy Transfer. Apoptosis induction by STS, H2O2 or selenite augmented the formation of VDAC oligomers several fold. The results show VDAC1 to be a component of the apoptosis machinery and offer new insight into the functions of VDAC1 oligomerization in apoptosis and of the VDAC1 N-terminal domain in the release of apoptogenic proteins as well as into regulation of VDAC by anti-apoptotic proteins, such as HK and Bcl2.

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    • "Apoptosis-mediated increase in VDAC1 oligomerization was observed regardless of the cell type or apoptosis inducer employed, including STS, curcumin, arsenic trioxide, etoposide, cisplatin, selenite, TNF-α, hydrogen peroxide, and UV light, all affecting mitochondria yet acting via different mechanisms. Conversely, the apoptosis inhibitor 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) prevented STSinduced VDAC1 oligomerization and apoptosis [25] [37]. At the same time, several studies demonstrated an increase in VDAC1 expression levels following apoptosis induction. "
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    ABSTRACT: VDAC1, an outer mitochondrial membrane (OMM) protein, is crucial for regulating mitochondrial metabolic and energetic functions and acts as a convergence point for various cell survival and death signals. VDAC1 is also a key player in apoptosis, involved in cytochrome c release and interactions with anti-apoptotic proteins. Recently, we demonstrated that various pro-apoptotic agents induce VDAC1 oligomerization and proposed that a channel formed by VDAC1 oligomers mediates cytochrome c release. As VDAC1 transports Ca(2+) across the OMM and because Ca(2+) has been implicated in apoptosis induction, we addressed the relationship between cytosolic Ca(2+) levels ([Ca2+]i), VDAC1 oligomerization and apoptosis induction. We demonstrate that different apoptosis inducers elevate cytosolic Ca(2+) and induce VDAC1 over-expression. Direct elevation of [Ca(2+)]i by the Ca(2+)-mobilizing agents A23187, ionomycin and thapsigargin also resulted in VDAC1 over-expression, VDAC1 oligomerization and apoptosis. In contrast, decreasing [Ca(2+)]i using the cell-permeable Ca(2+)-chelating reagent BAPTA-AM inhibited VDAC1 over-expression, VDAC1 oligomerization and apoptosis. Correlation between the increase in VDAC1 levels and oligomerization, [Ca(2+)]i levels and apoptosis induction, as induced by H2O2 or As2O3, was also obtained. On the other hand, cells transfected to overexpress VDAC1 presented Ca(2+)-independent VDAC1 oligomerization, cytochrome c release and apoptosis, suggesting that [Ca(2+)]i elevation is not pre-requisite for apoptosis induction when VDAC1 is over-expressed. The results suggest that Ca(2+) promotes VDAC1 over-expression by an as yet unknown signaling pathway, leading to VDAC1 oligomerization, ultimately resulting in apoptosis. These findings provide new insight into the mechanism of action of existing anti-cancer drugs involving induction of VDAC1 over-expression as a mechanism for inducing apoptosis. This article is part of a Special Issue entitled: Calcium Signaling In Health and Disease.
    Biochimica et Biophysica Acta (BBA) - Molecular Cell Research 04/2014; 1843(10). DOI:10.1016/j.bbamcr.2014.03.021 · 5.30 Impact Factor
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    • "RuR did not interact with E72Q-VDAC1 to reduce its channel activity or protect against apoptosis in cells expressing this mutant [17]. RuR-and AzRu-mediated protection against cell death, as induced by several apoptotic stimuli [24] [25] [26] [27] [28] [29], may arise from interaction with a VDAC1 Ca 2+ -binding site or with a specific protein conformation or by inhibiting mitochondrial Ca 2+ transport. These findings also indicate that VDAC1 functions as a Ca 2+ -sensitive Ca 2+ transporter in the OMM. "
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    ABSTRACT: The voltage-dependent anion channel (VDAC), located at the outer mitochondria membrane (OMM), mediates interactions between mitochondria and other parts of the cell by transporting anions, cations, ATP, Ca(2+), and metabolites. Substantial evidence points to VDAC1 as being a key player in apoptosis, regulating the release of apoptogenic proteins from mitochondria, such as cytochrome c, and interacting with anti-apoptotic proteins. Recently, we demonstrated that VDAC1 oligomerization is a general mechanism common to numerous apoptogens acting via different initiating cascades and proposed that a protein-conducting channel formed within a VDAC1 homo/hetro oligomer mediates cytochrome c release. However, the molecular mechanism responsible for VDAC1 oligomerization remains unclear. Several studies have shown that mitochondrial Ca(2+) is involved in apoptosis induction and that VDAC1 possesses Ca(2+)-binding sites and mediates Ca(2+) transport across the OMM. Here, the relationship between the cellular Ca(2+) level, [Ca(2+)]i, VDAC1 oligomerization and apoptosis was studied. Decreasing [Ca(2+)]i using the cell-permeable Ca(2+) chelating reagent BAPTA-AM was found to inhibit VDAC1 oligomerization and apoptosis, while increasing [Ca(2+)]i using Ca(2+) ionophore resulted in VDAC1 oligomerization and apoptosis induction in the absence of apoptotic stimuli. Moreover, induction of apoptosis elevated [Ca(2+)]i, concomitantly with VDAC1 oligomerization. AzRu-mediated inhibition of mitochondrial Ca(2+) transport decreased VDAC1 oligomerization, suggesting that mitochondrial Ca(2+) is required for VDAC1 oligomerization. In addition, increased [Ca(2+)]i levels up-regulate VDAC1 expression. These results suggest that Ca(2+) promotes VDAC1 oligomerization via activation of a yet unknown signaling pathway or by increasing VDAC1 expression, leading to apoptosis. This article is part of a Special Issue entitled: 12th European Symposium on Calcium.
    Biochimica et Biophysica Acta (BBA) - Molecular Cell Research 03/2013; DOI:10.1016/j.bbamcr.2013.03.017 · 5.30 Impact Factor
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    • "PTP, comprising of voltage-dependent anion channel (VDAC), adenine nucleotide translocase (ANT), and cyclophilin D, is located at contact sites between the inner and OMM [7], and PTP opening can be modulated by mitochondrial membrane potential (ΔΨ), elevated Ca 2+ , oxidative stress, thiol oxidation, or altered pyridine nucleotide status [9]. Since different apoptotic stimuli mediate VDAC oligomerization and channel formation, it is conceivable that a sufficiently large channel could allow mitochondria-to-cytosol translocation of apoptogenic factors [10] [11]. Moreover, OMM permeabilization and factor release could result from channel formation through Bax, Bak, and Bid conformational changes and homo-or hetero-oligomerization within the OMM [12]. "
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    ABSTRACT: Apoptosis is a highly organized form of cell death that is important for tissue homeostasis, organ development and senescence. To date, the extrinsic (death receptor mediated) and intrinsic (mitochondria derived) apoptotic pathways have been characterized in mammalian cells. Reduced glutathione, is the most prevalent cellular thiol that plays an essential role in preserving a reduced intracellular environment. glutathione protection of cellular macromolecules like deoxyribose nucleic acid proteins and lipids against oxidizing, environmental and cytotoxic agents, underscores its central anti-apoptotic function. Reactive oxygen and nitrogen species can oxidize cellular glutathione or induce its extracellular export leading to the loss of intracellular redox homeostasis and activation of the apoptotic signaling cascade. Recent evidence uncovered a novel role for glutathione involvement in apoptotic signaling pathways wherein post-translational S-glutathiolation of protein redox active cysteines is implicated in the potentiation of apoptosis. In the present review we focus on the key aspects of glutathione redox mechanisms associated with apoptotic signaling that includes: (a) changes in cellular glutathione redox homeostasis through glutathione oxidation or GSH transport in relation to the initiation or propagation of the apoptotic cascade, and (b) evidence for S-glutathiolation in protein modulation and apoptotic initiation.
    Biochimica et Biophysica Acta 06/2012; 1823(10):1767-77. DOI:10.1016/j.bbamcr.2012.06.019 · 4.66 Impact Factor
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