Loss of Drp1 function alters OPA1 processing and changes mitochondrial membrane organization.
ABSTRACT RNAi mediated loss of Drp1 function changes mitochondrial morphology in cultured HeLa and HUVEC cells by shifting the balance of mitochondrial fission and fusion towards unopposed fusion. Over time, inhibition of Drp1 expression results in the formation of a highly branched mitochondrial network along with "bulge"-like structures. These changes in mitochondrial morphology are accompanied by a reduction in levels of Mitofusin 1 (Mfn1) and 2 (Mfn2) and a modified proteolytic processing of OPA1 isoforms, resulting in the inhibition of cell proliferation. In addition, our data imply that bulge formation is driven by Mfn1 action along with particular proteolytic short-OPA1 (s-OPA1) variants: Loss of Mfn2 in the absence of Drp1 results in an increase of Mfn1 levels along with processed s-OPA1-isoforms, thereby enhancing continuous "fusion" and bulge formation. Moreover, bulge formation might reflect s-OPA1 mitochondrial membrane remodeling activity, resulting in the compartmentalization of cytochrome c deposits. The proteins Yme1L and PHB2 appeared not associated with the observed enhanced OPA1 proteolysis upon RNAi of Drp1, suggesting the existence of other OPA1 processing controlling proteins. Taken together, Drp1 appears to affect the activity of the mitochondrial fusion machinery by unbalancing the protein levels of mitofusins and OPA1.
- SourceAvailable from: Ana Raquel Esteves[Show abstract] [Hide abstract]
ABSTRACT: In Parkinson's disease mitochondrial dysfunction can lead to a deficient ATP supply to microtubule protein motors leading to mitochondrial axonal transport disruption. Compromised axonal transport will then lead to a disorganized distribution of mitochondria and other organelles in the cell, as well as, the accumulation of aggregated proteins like alpha-synuclein. Moreover, axonal transport disruption can trigger synaptic accumulation of autophagosomes packed with damaged mitochondria and protein aggregates promoting synaptic failure. We previously observed that neuronal-like cells with an inherent mitochondrial impairment derived from PD patients contain a disorganized microtubule network, as well as, alpha-synuclein oligomers accumulation. In this work we provide new evidence that an agent that promotes microtubule network assembly, NAP (davunetide), improves microtubule-dependent traffic, restores the autophagic flux and potentiates autophasosome-lysosome fusion leading to autophagic vacuole clearance in Parkinson's disease cells. Moreover, NAP is capable of efficiently reduce alpha-synuclein oligomer content and its sequestration by the mitochondria. Most interestingly, NAP decreases mitochondrial ubiquitination levels, as well as, increases mitochondrial membrane potential indicating a rescue in mitochondrial function. Overall, we demonstrate that by improving microtubule-mediated traffic, we can avoid mitochondrial-induced damage and thus recover cell homeostasis. These results prove that NAP may be a promising therapeutic lead candidate for neurodegenerative diseases that involve axonal transport failure and mitochondrial impairment as hallmarks, like Parkinson's disease and related disorders.Biochimica et Biophysica Acta 10/2013; · 4.66 Impact Factor
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ABSTRACT: Direct DNA damage is often considered the primary cause of cancer in patients exposed to ionizing radiation or environmental carcinogens. While mitochondria are known to play an important role in radiation-induced cellular response, the mechanisms by which cytoplasmic stimuli modulate mitochondrial dynamics and functions are largely unknown. In the present study, we examined changes in mitochondrial dynamics and functions triggered by α particle damage to the mitochondria in human small airway epithelial cells, using a precision microbeam irradiator with a beam width of one micron. Targeted cytoplasmic irradiation using this device resulted in mitochondrial fragmentation and a reduction of cytochrome c oxidase and succinate dehydrogenase activity, when compared with non-irradiated controls, suggesting a reduction in respiratory chain function. Additionally, mitochondrial fragmentation or fission was associated with increased expression of the dynamin-like protein DRP1, which promotes mitochondrial fission. DRP1 inhibition by the drug mdivi-1 prevented radiation-induced mitochondrial fission, but respiratory chain function in mitochondria inhibited by radiation persisted for 12 hr. Irradiated cells also showed an increase in mitochondria-derived superoxide that could be quenched by dimethyl sulfoxide. Taken together, our results provide a mechanistic explanation for the extranuclear, non-targeted effects of ionizing radiation.Cancer Research 09/2013; · 9.28 Impact Factor
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ABSTRACT: Mitochondria dysfunction is implicated in diverse conditions, including metabolic and neurodegenerative disorders. Mitochondrial dynamics has attracted increasing attention as to its relationship with mitochondria autophagy, also known as mitophagy, which is critical for degradation of dysfunctional mitochondria maintaining mitochondrial homeostasis. Mitochondrial ﬁssion and its role in clearance of injured mitochondria in acute ischemic injury, however, have not been elucidated yet. Here we showed that hypoxic/ischemic conditions led to fragmentation of mitochondria and induction of mitophagy in permanent middle cerebral artery occlusion (pMCAO) rats and oxygen-glucose deprivation (OGD) PC12 cells. Inhibition of Drp1 by pharmacologic inhibitor or siRNA resulted in accumulation of damaged mitochondria mainly through selectively blocking mitophagy without affecting mitochondrial biogenesis and non-selective autophagy. Drp1 inhibitors increased the infarct volume and aggravated the neurological deﬁcits in a rat model of pMCAO. We demonstrated that the devastating role of disturbed mitochondrial fission by inhibiting Drp1 contributed to the damaged mitochondria-mediated injury such as ROS generation, cyt-c release and activation of caspase-3. Taken together, we proved that under hypoxic/ischemic stress a Drp1-dependent mitophagy was triggered which was involved in the removal of damaged mitochondria and cellular survival at the early stage of hypoxic/ischemic injury. Thus, Drp1 related pathway involved in selective removal of dysfunctional mitochondria is proposed as an efficient target for treatment of cerebral ischemia.Neuropharmacology 07/2014; · 4.11 Impact Factor