Mitophagy is a highly specialized process to remove dysfunctional or superfluous mitochondria through the macroautophagy/autophagy pathway, aimed at protecting cells from the damage of disordered mitochondrial metabolism and apoptosis induction. PINK1, a neuroprotective protein mutated in autosomal recessive Parkinson disease, has been implicated in the activation of mitophagy by selectively accumulating on depolarized mitochondria, and promoting PARK2/Parkin translocation to them. While these steps have been characterized in depth, less is known about the process and site of autophagosome formation upon mitophagic stimuli. A previous study reported that, in starvation-induced autophagy, the proautophagic protein BECN1/Beclin1 (which we previously showed to interact with PINK1) relocalizes at specific regions of contact between the endoplasmic reticulum (ER) and mitochondria called mitochondria-associated membranes (MAM), from which the autophagosome originates. Here we show that, following mitophagic stimuli, autophagosomes also form at MAM; moreover, endogenous PINK1 and BECN1 were both found to relocalize at MAM, where they promoted the enhancement of ER-mitochondria contact sites and the formation of omegasomes, that represent autophagosome precursors. PARK2 was also enhanced at MAM following mitophagy induction. However, PINK1 silencing impaired BECN1 enrichment at MAM independently of PARK2, suggesting a novel role for PINK1 in regulating mitophagy. MAM have been recently implicated in many key cellular events. In this light, the observed prevalent localization of PINK1 at MAM may well explain other neuroprotective activities of this protein, such as modulation of mitochondrial calcium levels, mitochondrial dynamics, and apoptosis.

Mitochondria are considered as the power‐generating units of the cell due to their key role in energy metabolism and cell signaling. However, mitochondrial components could be found in the extracellular space, as fragments or encapsulated in vesicles. In addition, this intact organelle has been recently reported to be released by platelets exclusively in specific conditions. Here, we demonstrate for the first time, that blood preparation with resting platelets, contains whole functional mitochondria in normal physiological state. Likewise, we show, that normal and tumor cultured cells are able to secrete their mitochondria. Using serial centrifugation or filtration followed by polymerase chain reaction‐based methods, and Whole Genome Sequencing, we detect extracellular full‐length mitochondrial DNA in particles over 0.22 µm holding specific mitochondrial membrane proteins. We identify these particles as intact cell‐free mitochondria using fluorescence‐activated cell sorting analysis, fluorescence microscopy, and transmission electron microscopy. Oxygen consumption analysis revealed that these mitochondria are respiratory competent. In view of previously described mitochondrial potential in intercellular transfer, this discovery could greatly widen the scope of cell‐cell communication biology. Further steps should be developed to investigate the potential role of mitochondria as a signaling organelle outside the cell and to determine whether these circulating units could be relevant for early detection and prognosis of various diseases.

Mutations in the PINK1 gene cause autosomal recessive Parkinson's disease. The PINK1 gene encodes a protein kinase that is mitochondrially cleaved to generate two mature isoforms. In addition to its protective role against mitochondrial dysfunction and apoptosis, PINK1 is also known to regulate mitochondrial dynamics acting upstream of the PD-related protein Parkin. Recent data showed that mitochondrial Parkin promotes the autophagic degradation of dysfunctional mitochondria, and that stable PINK1 silencing may have an indirect role in mitophagy activation. Here we report a new interaction between PINK1 and Beclin1, a key pro-autophagic protein already implicated in the pathogenesis of Alzheimer's and Huntington's diseases. Both PINK1 N-and C-terminal are required for the interaction, suggesting that full-length PINK1, and not its cleaved isoforms, interacts with Beclin1. We also demonstrate that PINK1 significantly enhances basal and starvation-induced autophagy, which is reduced by knocking down Beclin1 expression or by inhibiting the Beclin1 partner Vps34. A mutant, PINK1 W437X , interaction of which with Beclin1 is largely impaired, lacks the ability to enhance autophagy, whereas this is not observed for PINK1 G309D , a mutant with defective kinase activity but unaltered ability to bind Beclin1. These findings identify a new function of PINK1 and further strengthen the link between autophagy and proteins implicated in the neurodegenerative process. Parkinson's disease (PD) is a frequent neurodegenerative disorder resulting from massive degeneration of the dopaminergic neurons in the substantia nigra. Although most cases are sporadic, several genes are known to cause familial PD, especially with early onset. 1 Mutations in the PINK1 gene are the second most frequent cause of autosomal recessive PD after those in the Parkin gene. 2,3 The PINK1 gene encodes a serine-threonine kinase with an N-terminal mitochondrial import sequence, first characterized as a protein aimed at maintaining mitochondrial integrity and preventing apoptosis in response to cellular stressors. 2,[4][5][6][7][8] This neuroprotective role is partly exerted through phosphorylation of the mitochondrial chaperon, TRAP1, although cytoplasm-restricted PINK1 was also shown to protect against MPTP damage. 9,10 The full-length PINK1 (PINK1-FL) is processed within mitochondria to generate two mature proteins; 4,11 all three isoforms localize both to the mitochondria and cytosol, their relative ratio being regulated by several factors. [10][11][12][13] Increasing data have demonstrated that absence of functional PINK1 induces abnormalities of mitochondrial morphology. 6,14,15 In several studies (mostly in Drosophila), PINK1 was shown to promote fission acting upstream of the Fis1-Drp1 machinery, and the mitochondrial phenotype observed in PINK1 knockout flies or silenced cells was associated to reduced fission. 16,17 Subsequent studies in mammalian cell systems contradicted these results, demonstrating that mutant or silenced PINK1 resulted in incre...

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Mutations in the PINK1 gene are a frequent cause of autosomal recessive Parkinson's disease (PD). PINK1 encodes a mitochondrial kinase with neuroprotective activity, implicated in maintaining mitochondrial homeostasis and function. In concurrence with Parkin, PINK1 regulates mitochondrial trafficking and degradation of damaged mitochondria through mitophagy. Moreover, PINK1 can activate autophagy by interacting with the pro-autophagic protein Beclin-1. Here, we report that, upon mitochondrial depolarization, PINK1 interacts with and phosphorylates Bcl-xL, an anti-apoptotic protein also known to inhibit autophagy through its binding to Beclin-1. PINK1-Bcl-xL interaction does not interfere either with Beclin-1 release from Bcl-xL or the mitophagy pathway; rather it protects against cell death by hindering the pro-apoptotic cleavage of Bcl-xL. Our data provide a functional link between PINK1, Bcl-xL and apoptosis, suggesting a novel mechanism through which PINK1 regulates cell survival. This pathway could be relevant for the pathogenesis of PD as well as other diseases including cancer. Parkinson's disease (PD) is the second most common neurodegenerative disorder after Alzheimer's disease, with prevalence of 1% in the population older than 60 years. 1 Several biochemical abnormalities, including mitochondrial dysfunction, oxidative stress and misfolded protein damage, have been implicated in PD pathogenesis. 2 Although the majority of late-onset cases are sporadic, early-onset PD is frequently caused by mutations in genes with autosomal recessive inheritance, mainly Parkin (GeneID: 5071) and PINK1 (GeneID: 65018). 3 PINK1 encodes a 63 kDa mitochondrial protein kinase, which is processed by mitochondrial proteases to generate two smaller isoforms. [4][5][6][7] We and others have shown that PINK1 acts as a key neuroprotective protein, aimed at preventing mitochondrial dysfunction and apoptotic cell death in response to multiple stress conditions. [8][9][10] This pro-survival activity is exerted through several mechanisms, including phosphorylation of the mitochondrial proteins TRAP1 and Omi/HtrA2, and regulation of mitochondrial calcium buffering. [11][12][13][14] Increasing data now indicate that PINK1 acts upstream of Parkin in an evolutionary conserved pathway implicated in regulating mitochondrial biogenesis, trafficking and fusion/ fission events, to maintain mitochondrial network health. 15 In particular, upon mitochondrial depolarization, PINK1 processing is impaired, determining a marked accumulation of the full-length protein on the surface of dysfunctional mitochondria, where it recruits Parkin. This process results in the phosphorylation and/or ubiquitination of several mitochondrial substrates, leading to the selective quarantine of damaged mitochondria and their degradation through mitophagy. [16][17][18][19] In line with this, we reported that coexpression of mutant, but not wild-type (wt) PINK1, with mutant alpha-synuclein resulted in the formation of enlarged autophagosomes surrounding abnormal mitoch...

The mouse double minute 2 (MDM2) oncoprotein is recognized as a major negative regulator of the p53 tumor suppressor, but growing evidence indicates that its oncogenic activities extend beyond p53. Here, we show that MDM2 is recruited to chromatin independently of p53 to regulate a transcriptional program implicated in amino acid metabolism and redox homeostasis. Identification of MDM2 target genes at the whole-genome level highlights an important role for ATF3/4 transcription factors in tethering MDM2 to chromatin. MDM2 recruitment to chromatin is a tightly regulated process that occurs during oxidative stress and serine/glycine deprivation and is modulated by the pyruvate kinase M2 (PKM2) metabolic enzyme. Depletion of endogenous MDM2 in p53-deficient cells impairs serine/glycine metabolism, the NAD(+)/NADH ratio, and glutathione (GSH) recycling, impacting their redox state and tumorigenic potential. Collectively, our data illustrate a previously unsuspected function of chromatin-bound MDM2 in cancer cell metabolism.

Accumulating evidence indicates that the MDM2 oncoprotein promotes tumorigenesis beyond its canonical negative effects on the p53 tumor suppressor, but these p53-independent functions remain poorly understood. Here, we show that a fraction of endogenous MDM2 is actively imported in mitochondria to control respiration and mitochondrial dynamics independently of p53. Mitochondrial MDM2 represses the transcription of NADH-dehydrogenase 6 (MT-ND6) in vitro and in vivo, impinging on respiratory complex I activity and enhancing mitochondrial ROS production. Recruitment of MDM2 to mitochondria increases during oxidative stress and hypoxia. Accordingly, mice lacking MDM2 in skeletal muscles exhibit higher MT-ND6 levels, enhanced complex I activity, and increased muscular endurance in mild hypoxic conditions. Furthermore, increased mitochondrial MDM2 levels enhance the migratory and invasive properties of cancer cells. Collectively, these data uncover a previously unsuspected function of the MDM2 oncoprotein in mitochondria that play critical roles in skeletal muscle physiology and may contribute to tumor progression.

The gene PINK1 [phosphatase and tensin homologue (PTEN)-induced putative kinase 1] encodes a serine/threonine kinase which was initially linked to the pathogenesis of a familial form of Parkinson's disease. Research on PINK1 has recently unravelled that its multiple functions extend well beyond neuroprotection, implicating this eclectic protein in a growing number of human pathologies, including cancer, diabetes, cardiopulmonary dysfunctions, and inflammation. Extensive studies have identified PINK1 as a crucial player in the mitochondrial quality control pathway, required to label damaged mitochondria and promote their elimination through an autophagic process (mitophagy). Mounting evidence now indicates that PINK1 activities are not restricted solely to mitophagy, and that different subcellular and even sub-mitochondrial pools of PINK1 are involved in distinct signalling cascades to regulate cell metabolism and survival. In this review, we provide a concise overview on the different functions of PINK1 and their potential role in human diseases.