VDAC1 at the crossroads of cell metabolism, apoptosis and cell stress.
Ontology highlight
ABSTRACT:
SUBMITTER: Shoshan-Barmatz V
PROVIDER: S-EPMC6287957 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Carcinogenesis is a complicated process that involves the deregulation of epigenetics, resulting in cellular transformational events, such as proliferation, differentiation, and metastasis. Most chromatin-modifying enzymes utilize metabolites as co-factors or substrates and thus are directly dependent on such metabolites as acetyl-coenzyme A, S-adenosylmethionine, and NAD+. Here, we show that using specific siRNA to deplete a tumor of VDAC1 not only led to reprograming of the cancer cell metabolism but also altered several epigenetic-related enzymes and factors. VDAC1, in the outer mitochondrial membrane, controls metabolic cross-talk between the mitochondria and the rest of the cell, thus regulating the metabolic and energetic functions of mitochondria, and has been implicated in apoptotic-relevant events. We previously demonstrated that silencing VDAC1 expression in glioblastoma (GBM) U-87MG cell-derived tumors, resulted in reprogramed metabolism leading to inhibited tumor growth, angiogenesis, epithelial-mesenchymal transition and invasiveness, and elimination of cancer stem cells, while promoting the differentiation of residual tumor cells into neuronal-like cells. These VDAC1 depletion-mediated effects involved alterations in transcription factors regulating signaling pathways associated with cancer hallmarks. As the epigenome is sensitive to cellular metabolism, this study was designed to assess whether depleting VDAC1 affects the metabolism-epigenetics axis. Using DNA microarrays, q-PCR, and specific antibodies, we analyzed the effects of si-VDAC1 treatment of U-87MG-derived tumors on histone modifications and epigenetic-related enzyme expression levels, as well as the methylation and acetylation state, to uncover any alterations in epigenetic properties. Our results demonstrate that metabolic rewiring of GBM via VDAC1 depletion affects epigenetic modifications, and strongly support the presence of an interplay between metabolism and epigenetics.
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Project description:Background/aimsThe role of VDAC1, the most abundant mitochondrial outer membrane protein, in cell death depends on cell types and stimuli. Both silencing and upregulation of VDAC1 in various type of cancer cell lines can stimulate apoptosis. In contrast, in mouse embryonic stem (MES) cells and mouse embryonic fibroblasts (MEFs), the roles of VDAC1 knockout (VDAC1-/-) in apoptotic cell death are contradictory. The contribution and underlying mechanism of VDAC1-/- in oxidative stress-induced cell death in cardiac cells has not been established. We hypothesized that VDAC1 is an essential regulator of oxidative stress-induced cell death in H9c2 cells.MethodsWe knocked out VDAC1 in this rat cardiomyoblast cell line with CRISPR-Cas9 genome editing technique to produce VDAC1-/- H9c2 cells, and determined if VDAC1 is critical in promoting cell death via oxidative stress induced by tert-butylhydroperoxide (tBHP), an organic peroxide, or rotenone (ROT), an inhibitor of mitochondrial complex I by measuring cell viability with MTT assay, cell death with TUNEL stain and LDH release. The mitochondrial and glycolytic stress were examined by measuring O2 consumption rate (OCR) and extracellular acidification rate (ECAR) with a Seahorse XFp analyzer.ResultsWe found that under control conditions, VDAC1-/- did not affect H9c2 cell proliferation or mitochondrial respiration. However, compared to the wildtype (WT) cells, exposure to either tBHP or ROT enhanced the production of ROS, ECAR, and the proton (H+) production rate (PPR) from glycolysis, as well as promoted apoptotic cell death in VDAC1-/- H9c2 cells. VDAC1-/- H9c2 cells also exhibited markedly reduced mitochondria-bound hexokinase II (HKII) and Bax. Restoration of VDAC1 in VDAC1-/- H9c2 cells reinstated mitochondria-bound HKII and concomitantly decreased tBHP and ROT-induced ROS production and cell death. Interestingly, mitochondrial respiration remained the same after tBHP treatment in VDAC1-/- and WT H9c2 cells.ConclusionOur results suggest that VDAC1-/- in H9c2 cells enhances oxidative stress-mediated cell apoptosis that is directly linked to the reduction of mitochondria-bound HKII and concomitantly associated with enhanced ROS production, ECAR, and PPR.
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Project description:VDAC1 is a critical substrate of Parkin responsible for the regulation of mitophagy and apoptosis. Here, we demonstrate that VDAC1 can be either mono- or polyubiquitinated by Parkin in a PINK1-dependent manner. VDAC1 deficient with polyubiquitination (VDAC1 Poly-KR) hampers mitophagy, but VDAC1 deficient with monoubiquitination (VDAC1 K274R) promotes apoptosis by augmenting the mitochondrial calcium uptake through the mitochondrial calcium uniporter (MCU) channel. The transgenic flies expressing Drosophila Porin K273R, corresponding to human VDAC1 K274R, show Parkinson disease (PD)-related phenotypes including locomotive dysfunction and degenerated dopaminergic neurons, which are relieved by suppressing MCU and mitochondrial calcium uptake. To further confirm the relevance of our findings in PD, we identify a missense mutation of Parkin discovered in PD patients, T415N, which lacks the ability to induce VDAC1 monoubiquitination but still maintains polyubiquitination. Interestingly, Drosophila Parkin T433N, corresponding to human Parkin T415N, fails to rescue the PD-related phenotypes of Parkin-null flies. Taken together, our results suggest that VDAC1 monoubiquitination plays important roles in the pathologies of PD by controlling apoptosis.
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2011-06-01 | GSE24065 | GEO
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