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Glutathionylated and Fe-S cluster containing hMIA40 (CHCHD4) regulates ROS and mitochondrial complex III and IV activities of the electron transport chain.

ABSTRACT: Human MIA40, an intermembrane space (IMS) import receptor of mitochondria harbors twin CX9C motifs for stability while its CPC motif is known to facilitate the import of IMS bound proteins. Site-directed mutagenesis complemented by MALDI on in vivo hMIA40 protein shows that a portion of MIA40 undergoes reversible S-glutathionylation at three cysteines in the twin CX9C motifs and the lone cysteine 4 residue. We find that HEK293T cells expressing hMIA40 mutant defective for glutathionylation are compromised in the activities of complexes III and IV of the Electron Transport Chain (ETC) and enhance Reactive Oxygen Species (ROS) levels. Immunocapture studies show MIA40 interacting with complex III. Interestingly, glutathionylated MIA40 can transfer electrons to cytochrome C directly. However, Fe-S clusters associated with the CPC motif are essential to facilitate the two-electron to one-electron transfer for reducing cytochrome C. These results suggest that hMIA40 undergoes glutathionylation to maintain ROS levels and for optimum function of complexes III and IV of ETC. Our studies shed light on a novel post-translational modification of hMIA40 and its ability to act as a redox switch to regulate the ETC and cellular redox homeostasis.

SUBMITTER: Thiriveedi VR

PROVIDER: S-EPMC7511737 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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Glutathionylated and Fe-S cluster containing hMIA40 (CHCHD4) regulates ROS and mitochondrial complex III and IV activities of the electron transport chain.

Thiriveedi Venkata Ramana VR Mattam Ushodaya U Pattabhi Prasad P Bisoyi Vandana V Talari Noble Kumar NK Krishnamoorthy Thanuja T Sepuri Naresh Babu V NBV

Redox biology 20200915

Human MIA40, an intermembrane space (IMS) import receptor of mitochondria harbors twin CX9C motifs for stability while its CPC motif is known to facilitate the import of IMS bound proteins. Site-directed mutagenesis complemented by MALDI on in vivo hMIA40 protein shows that a portion of MIA40 undergoes reversible S-glutathionylation at three cysteines in the twin CX9C motifs and the lone cysteine 4 residue. We find that HEK293T cells expressing hMIA40 mutant defective for glutathionylation are c ...[more]

PMID: 32971361

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Project description:Loss of dopamine (DA) homeostasis may be a contributing factor to cell damage in Parkinson's disease (PD). Past studies showing deleterious effects of DA on mitochondrial function, however, have been inconsistent raising questions about mitochondria as a downstream target for DA. Issues such as the dopamine species i.e., reduced or oxidized, time of exposure and the effect of major metabolites such as 3,4-dihydrophenylacetic acid (DOPAC) may contribute to the disparate findings. The present study used isolated, lysed rat brain mitochondria to characterize the effects of oxidized or reduced DA and DOPAC on complex activities of the electron transport chain (ETC). Time of exposure and quantitation of reduced or oxidized catachols for DA and DOPAC were monitored for all experiments. Reduced DA and DOPAC with or without a 30min preincubation had no affect on NADH oxidase activity which monitors the activities of complexes I, III and IV. Complex II activity was inhibited by reduced DA (?500?M), but not by reduced DOPAC and was significantly attenuated by SOD suggesting reactive oxygen species involvement. In contrast, fully oxidized DA and DOPAC dose dependently inhibited NADH oxidase, complex I and complex III activities with IC(50s) in the 50-200?M range. No preincubation was required for inhibition with the catechols when they were fully oxidized. Oxidized DA inhibited complex I only when exposure occurred during stimulated electron flow, suggesting covalent binding of quinones to proteins within active sites of the complex. In intact, well coupled mitochondria, extramitochondrial DA was shown to access the mitochondrial matrix in a dose, time and energy-dependent fashion. The findings suggest that many of the reported inconsistencies with regards to the effects of DA and DOPAC on ETC function can be attributed to the oxidized state of the catechol at the time of exposure. In addition, the findings provide possible downstream targets for DA that could contribute to the vulnerability of dopaminergic neurons in PD.

Dataset Information

Glutathionylated and Fe-S cluster containing hMIA40 (CHCHD4) regulates ROS and mitochondrial complex III and IV activities of the electron transport chain.

Publications

Glutathionylated and Fe-S cluster containing hMIA40 (CHCHD4) regulates ROS and mitochondrial complex III and IV activities of the electron transport chain.

Similar Datasets

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