Human VDAC isoforms differ in their capability to interact with minocycline and to contribute to its cytoprotective activity

Karachitos, Andonis; Grobys, Daria; Antoniewicz, Monika; Jedut, Sylwia; Jordán, Joaquı́n; Kmita, Hanna

doi:10.1016/j.mito.2016.03.004

Cited by 11 publications

(14 citation statements)

References 71 publications

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“…In general, yeast or mammalian VDAC1 persists at low potentials in a stable highly conductive state, called the 'open' state. However, as the applied voltage rises (starting from AE20-30 mV), VDAC1 switches to several lowconducting states, called 'closed' states [24][25][26][27]. We demonstrated previously that the native yVDAC2 shows a less pronounced voltage dependence in comparison to yVDAC1, because yVDAC2-formed channel closure began from AE40-50 mV of applied voltage [18].…”

Section: Reconstituted Recombinant Yvdac2 Forms Channels Of Conductanmentioning

confidence: 95%

Recombinant yeast VDAC2: a comparison of electrophysiological features with the native form

et al. 2019

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Voltage‐dependent anion channel isoform 2 of the yeast Saccharomyces cerevisiae ( yVDAC 2) was believed for many years to be devoid of channel activity. Recently, we isolated yVDAC 2 and showed that it exhibits channel‐forming activity in the planar lipid bilayer system when in its so‐called native form. Here, we describe an alternative strategy for yVDAC 2 isolation, through heterologous expression in bacteria and refolding in vitro . Recombinant yVDAC 2, like its native form, is able to form voltage‐dependent channels. However, some differences between native and recombinant yVDAC 2 emerged in terms of voltage dependence and ion selectivity, suggesting that, in this specific case, the recombinant protein might be depleted of post‐translational modification(s) that occur in eukaryotic cells.

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Section: Reconstituted Recombinant Yvdac2 Forms Channels Of Conductanmentioning

confidence: 95%

Recombinant yeast VDAC2: a comparison of electrophysiological features with the native form

et al. 2019

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“…The status of mitochondrial coupling in intact cells can be estimated by calculation of mitochondrial coupling efficiency and mitochondrial coupling capacity. As mentioned in Section “Materials and Methods,” they correspond to the state 3 contribution to basal respiration and FCCP uncoupling capacity (the state U to state 4 ratio), respectively (29, 31). State 3 can be calculated by subtracting state 4 from basal respiration, and state 4 can be enforced by inhibitors of adenine nucleotide translocase or ATP synthase able to cross the cell membrane (i.e., by elimination of state 3), whereas state U (uncoupled state that connotes a maximal rate of oxygen uptake) can be induced by FCCP, which collapses the inner membrane potential.…”

Section: Resultsmentioning

confidence: 99%

“…It has been shown recently that all mammalian VDAC isoforms are able to form channels of comparable characteristic (31, 46, 47). Therefore, it may be speculated that dynamic movement of N-terminal helix defined as the vital element of the voltage-dependence and resulting conductance control [e.g., De Pinto et al (48)] is impaired in all VDAC isoforms due to the protein modification or interaction with mHtt.…”

Section: Discussionmentioning

confidence: 99%

“…State 4 (oligomycin-resistant respiration) was enforced by addition of 2 μg/ml oligomycin (an inhibitor of mitochondrial ATPase), whereas state U (uncoupled state) was induced by FCCP (carbonyl cyanide p-trifluoromethoxyphenylhydrazone; an uncoupler) added in portions of 0.1 μM each to avoid respiration inhibition (Figure 2A). To estimate the status of mitochondrial coupling in intact cells, basal respiration, contribution of state 3 to basal respiration, and uncoupling capacity of FCCP were calculated (29, 31). The state 3 (oligomycin-inhabitable respiration) was calculated by subtracting state 4 from basal respiration.…”

Section: Methodsmentioning

confidence: 99%

“…Artificial phospholipid membranes were formed from soybean asolectin (Avanti Polar Lipids, purified by acetone precipitation before use) suspended at concentration of 25 mg/ml in n-decane, across a circular hole (aperture diameter about 250 μm) in the thin wall of Delrin chamber (Warner Instruments) separating two compartments (cis–trans) filled with unbuffered 1M KCl (31, 34, 35). The pH of KCl solution was 6.9.…”

Section: Methodsmentioning

confidence: 99%

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The Association of VDAC with Cell Viability of PC12 Model of Huntington’s Disease

et al. 2016

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It is becoming increasingly apparent that mitochondria dysfunction plays an important role in the pathogenesis of Huntington’s disease (HD), but the underlying mechanism is still elusive. Thus, there is a still need for further studies concerning the upstream events in the mitochondria dysfunction that could contribute to cell death observed in HD. Taking into account the fundamental role of the voltage-dependent anion-selective channel (VDAC) in mitochondria functioning, it is reasonable to consider the channel as a crucial element in HD etiology. Therefore, we applied inducible PC12 cell model of HD to determine the relationship between the effect of expression of wild type and mutant huntingtin (Htt and mHtt, respectively) on cell survival and mitochondria functioning in intact cells under conditions of undergoing cell divisions. Because after 48 h of Htt and mHtt expression differences in mitochondria functioning co-occurred with differences in the cell viability, we decided to estimate the effect of Htt and mHtt expression lasted for 48 h on VDAC functioning. Therefore, we isolated VDAC from the cells and tested the preparations by black lipid membrane system. We observed that the expression of mHtt, but not Htt, resulted in changes of the open state conductance and voltage-dependence when compared to control cells cultured in the absence of the expression. Importantly, for all the VDAC preparations, we observed a dominant quantitative content of VDAC1, and the quantitative relationships between VDAC isoforms were not changed by Htt and mHtt expression. Thus, Htt and mHtt-mediated functional changes of VDAC, being predominantly VDAC1, which occur shortly after these protein appearances in cells, may result in differences concerning mitochondria functioning and viability of cells expressing Htt and mHtt. The assumption is important for better understanding of cytotoxicity as well as cytoprotection mechanisms of potential clinical application.

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yVDAC2, the second mitochondrial porin isoform of Saccharomyces cerevisiae

Guardiani¹,

Magrì²,

Karachitos³

et al. 2018

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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The yeast Saccharomyces cerevisiae genome is endowed with two distinct isoforms of Voltage-Dependent Anion Channel (VDAC). The isoform yVDAC2 is currently understudied with respect to the best known yVDAC1. Yet, since the discovery, the function of yVDAC2 was unclear, leading to the hypothesis that it might be devoid of a channel function. In this work we have elucidated, by bioinformatics modeling and electrophysiological analysis, the functional activity of yVDAC2. The conformation of yVDAC2 and, for comparison, of yVDAC1 were modeled using a multiple template approach involving mouse, human and zebrafish structures and both showed to arrange the sequences as the typical 19-stranded VDAC β-barrel. Molecular dynamics simulations showed that yVDAC2, in comparison with yVDAC1, has a different number of permeation paths of potassium and chloride ions. yVDAC2 protein was over-expressed in the S. cerevisiae cells depleted of functional yVDAC1 (Δpor1 mutant) and, after purification, it was reconstituted in artificial membranes (planar lipid bilayer (PLB) system). The protein displayed channel-forming activity and the calculated conductance, voltage-dependence and ion selectivity values were similar to those of yVDAC1 and other members of VDAC family. This is the first time that yVDAC2 channel features are detected and characterized.

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Human VDAC isoforms differ in their capability to interact with minocycline and to contribute to its cytoprotective activity

Cited by 11 publications

References 71 publications

Recombinant yeast VDAC2: a comparison of electrophysiological features with the native form

Recombinant yeast VDAC2: a comparison of electrophysiological features with the native form

The Association of VDAC with Cell Viability of PC12 Model of Huntington’s Disease

yVDAC2, the second mitochondrial porin isoform of Saccharomyces cerevisiae

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