Mediation of the Antiapoptotic Activity of Bcl-xL Protein upon Interaction with VDAC1 Protein

Department of Life Sciences and the National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel.
Journal of Biological Chemistry (Impact Factor: 4.57). 05/2012; 287(27):23152-61. DOI: 10.1074/jbc.M112.345918
Source: PubMed


The mitochondrial protein, the voltage-dependent anion channel (VDAC), is implicated in the control of apoptosis, including via its interaction with the pro- and antiapoptotic proteins. We previously demonstrated the direct interaction of Bcl2 with VDAC, leading to reduced channel conductance. VDAC1-based peptides interacted with Bcl2 to prevent its antiapoptotic activity. Here, using a variety of approaches, we show the interaction of the antiapoptotic protein, Bcl-xL, with VDAC1 and reveal that this interaction mediates Bcl-xL protection against apoptosis. C-terminally truncated Bcl-xL(Δ21) interacts with purified VDAC1, as revealed by microscale thermophoresis and as reflected in the reduced channel conductivity of bilayer-reconstituted VDAC1. Overexpression of Bcl-xL prevented staurosporine-induced apoptosis in cells expressing native VDAC1 but not certain VDAC1 mutants. Having identified mutations in VDAC1 that interfere with the Bcl-xL interaction, certain peptides representing VDAC1 sequences, including the N-terminal domain, were designed and generated as recombinant and synthetic peptides. The VDAC1 N-terminal region and two internal sequences were found to bind specifically, and in a concentration- and time-dependent manner, to immobilized Bcl-xL(Δ21), as revealed by surface plasmon resonance. Moreover, expression of the recombinant peptides in cells overexpressing Bcl-xL prevented protection offered by the protein against staurosporine-induced apoptosis. These results point to Bcl-xL acting as antiapoptotic protein, promoting tumor cell survival via binding to VDAC1. These findings suggest that interfering with Bcl-xL binding to the mitochondria by VDAC1-based peptides may serve to induce apoptosis in cancer cells and to potentiate the efficacy of conventional chemotherapeutic agents.

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    • "Cytochrome c activates caspases which eventually leads to cell death. Anti-apoptotic Bcl-2 proteins maintain mitochondrial integrity by sequestering pro-apoptotic members (Arbel, Ben-Hail, & Shoshan-Barmatz, 2012; Kinnally, Peixoto, Ryu, & Dejean, 2011; Shore, 2009). Glycolytic enzyme hexokinase (HK) interacts with VDAC, and this interaction makes the cells resistant to several apoptotic stimuli. "
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    ABSTRACT: Centella asiatica (CA) is commonly used as a leafy vegetable in many Asian countries. Consumption of CA is believed to prevent neuronal damage and improve brain function. The protective effect of CA on N2a cells were evaluated using ischaemia-reperfusion (IR) injury and oxygen-glucose deprivation (OGD) models in order to shed light on its molecular mechanism of action. Aqueous-methanolic extract of the CA leaves protected N2a cells against IR injury. CA reduced the levels of intracellular reactive oxygen species (ROS). It also prevented the elevation of intracellular calcium and attenuated the change of mitochondrial membrane potential, caused by OGD. When VDAC-1 was knocked down in N2a cells, CA failed to protect cells against IR injury. Further, CA modulated the properties of human VDAC-1 (hVDAC-1). hVDAC-1, when reconstituted in the lipid bilayer membrane, showed higher conductance after treating with CA. CA stabilized hVDAC-1 in open state, which is possibly associated with its cytoprotective effect.
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    • "For instance, many experiments performed by White et al. were based on mitochondrial Ca 2+ uptake experiments in permeabilized cells where all VDAC1 channels across the mitochondrial outer membrane will participate in mitochondrial Ca 2+ uptake[114]. In contrast, experiments in other studies relied on mitochondrial Ca 2+ uptake measurements in intact cells exposed to agonists for which Ca 2+ would be preferentially transferred via ER-mitochondrial contact sites[113,115,116]. Nevertheless, since a stimulatory effect of VDAC1 by Bcl-XL has never been shown in direct measurements based on purified VDAC1 channels, the molecular properties and the relevance of the VDAC1/ Bcl-XL connection in promoting mitochondrial Ca 2+ transfer ought to be further scrutinized. "
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    ABSTRACT: Intracellular Ca2 + signals that arise from the endoplasmic reticulum (ER), the major intracellular Ca2 +-storage organelle, impact several mitochondrial functions and dictate cell survival and cell death processes. Furthermore, alterations in Ca2 + signaling in cancer cells promote survival and establish a high tolerance towards cell stress and damage, so that the on-going oncogenic stress does not result in the activation of cell death. Over the last years, the mechanisms underlying these oncogenic alterations in Ca2 + signaling have started to emerge. An important aspect of this is the identification of several major oncogenes, including Bcl-2, Bcl-XL, Mcl-1, PKB/Akt, and Ras, and tumor suppressors, such as p53, PTEN, PML, BRCA1, and Beclin 1, as direct and critical regulators of Ca2 +-transport systems located at the ER membranes, including IP3 receptors and SERCA Ca2 + pumps. In this way, these proteins execute part of their function by controlling the ER-mitochondrial Ca2 + fluxes, favoring either survival (oncogenes) or cell death (tumor suppressors). Oncogenic mutations, gene deletions or amplifications alter the expression and/or function of these proteins, thereby changing the delicate balance between oncogenes and tumor suppressors, impacting oncogenesis and favoring malignant cell function and behavior. In this review, we provided an integrated overview of the impact of the major oncogenes and tumor suppressors, often altered in cancer cells, on Ca2 + signaling from the ER Ca2 + stores. This article is part of a Special Issue entitled: Calcium and Cell Fate. Guest Editors: Dr. Jacques Haiech, Professor Heizmann, Professor Joachim Krebs.
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    • "mitochondrial membrane, VDACs interact directly with a plethora of pro-and anti-apoptotic factors, either members of the Bcl-2 protein family such as Bcl-xL (Arbel et al., 2012; Malia and Wagner, 2007), Bid/tBid(Rostovtseva et al., 2004), Bax/Bak, (Shimizu et al., 2001), hexokinases (Godbole et al., 2013) or Bnip3 (Chaanine et al., 2013). The precise molecular functions and stoichiometries of these VDAC/apoptotic factor complexes are poorly understood, although tentative structural predictions for several VDAC/effector complexes have been suggested (G.Veresov and Davidovskii, 2014). "
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