Mitochondrial-derived Vesicles Research Articles

Mitolysosome exocytosis: a novel mitochondrial quality control pathway linked with parkinsonism-like symptoms.

Quality control of mitochondria is essential for their homeostasis and function. Light chain 3 (LC3) associated autophagosomes-mediated mitophagy represents a canonical mitochondrial quality control pathway. Alternative quality control processes, such as mitochondrial-derived vesicles (MDVs), have been discovered, but the intact mitochondrial quality control remains unknown. We recently discovered a novel mitolysosome exocytosis mechanism for mitochondrial quality control in flunarizine (FNZ)-induced mitochondria clearance, where autophagosomes are not required, but rather mitochondria are engulfed directly by lysosomes, mediating mitochondrial secretion. As FNZ results in parkinsonism, we propose that excessive mitolysosome exocytosis is the cause.

Mitochondrial Damage-Associated Molecular Patterns Content in Extracellular Vesicles Promotes Early Inflammation in Neurodegenerative Disorders.

Neuroinflammation is a common hallmark in different neurodegenerative conditions that share neuronal dysfunction and a progressive loss of a selectively vulnerable brain cell population. Alongside ageing and genetics, inflammation, oxidative stress and mitochondrial dysfunction are considered key risk factors. Microglia are considered immune sentinels of the central nervous system capable of initiating an innate and adaptive immune response. Nevertheless, the pathological mechanisms underlying the initiation and spread of inflammation in the brain are still poorly described. Recently, a new mechanism of intercellular signalling mediated by small extracellular vesicles (EVs) has been identified. EVs are nanosized particles (30–150 nm) with a bilipid membrane that carries cell-specific bioactive cargos that participate in physiological or pathological processes. Damage-associated molecular patterns (DAMPs) are cellular components recognised by the immune receptors of microglia, inducing or aggravating neuroinflammation in neurodegenerative disorders. Diverse evidence links mitochondrial dysfunction and inflammation mediated by mitochondrial-DAMPs (mtDAMPs) such as mitochondrial DNA, mitochondrial transcription factor A (TFAM) and cardiolipin, among others. Mitochondrial-derived vesicles (MDVs) are a subtype of EVs produced after mild damage to mitochondria and, upon fusion with multivesicular bodies are released as EVs to the extracellular space. MDVs are particularly enriched in mtDAMPs which can induce an immune response and the release of pro-inflammatory cytokines. Importantly, growing evidence supports the association between mitochondrial dysfunction, EV release and inflammation. Here, we describe the role of extracellular vesicles-associated mtDAMPS in physiological conditions and as neuroinflammation activators contributing to neurodegenerative disorders.

Matrix Vesicle–Mediated Mineralization and Potential Applications

Hard tissues, including the bones and teeth, are a fundamental part of the body, and their formation and homeostasis are critically regulated by matrix vesicle–mediated mineralization. Matrix vesicles have been studied for 50 y since they were first observed using electron microscopy. However, research progress has been hampered by various technical barriers. Recently, there have been great advancements in our understanding of the intracellular biosynthesis of matrix vesicles. Mitochondria and lysosomes are now considered key players in matrix vesicle formation. The involvement of mitophagy, mitochondrial-derived vesicles, and mitochondria–lysosome interaction have been suggested as potential detailed mechanisms of the intracellular pathway of matrix vesicles. Their main secretion pathway may be exocytosis, in addition to the traditionally understood mechanism of budding from the outer plasma membrane. This basic knowledge of matrix vesicles should be strengthened by novel nano-level microscopic technologies, together with basic cell biologies, such as autophagy and interorganelle interactions. In the field of tissue regeneration, extracellular vesicles such as exosomes are gaining interest as promising tools in cell-free bone and periodontal regenerative therapy. Matrix vesicles, which are recognized as a special type of extracellular vesicles, could be another potential alternative. In this review, we outline the recent significant progress in the process of matrix vesicle–mediated mineralization and the potential clinical applications of matrix vesicles for tissue regeneration.

Mitochondrial-derived vesicles: Recent insights.

The generation of vesicles is a constitutive attribute of mitochondria inherited from bacterial ancestors. The physiological conditions and mild oxidative stress promote oxidation and dysfunction of certain proteins and lipids within the mitochondrial membranes; these constituents are subsequently packed as small mitochondrial‐derived vesicles (MDVs) (70–150 nm in diameter) and are transported intracellularly to lysosomes and peroxisomes to be degraded. In this way, MDVs remove the damaged mitochondrial components, preserve mitochondrial structural and functional integrity and restore homeostasis. An outline of the current knowledge on MDVs seems to be necessary for understanding the potential impact of this research area in cellular (patho)physiology. The present synopsis is an attempt towards the accomplishment of this demand, highlighting also the still unclear issues related to MDVs. Here, we discuss (i) MDVs budding and generation (molecules and mechanisms), (ii) the distinct cargoes packed and transported by MDVs, (iii) the MDVs trafficking pathways and (iv) the biological role of MDVs, from quality controllers to the involvement in organellar crosstalk, mitochondrial antigen presentation and peroxisome de novo biogenesis. These complex roles uncover also mitochondria integration into the cellular environment. As the therapeutic exploitation of MDVs is currently limited, future insights into MDVs cell biology are expected to direct to novel diagnostic tools and treatments.

Exosomes in Parkinson: Revisiting Their Pathologic Role and Potential Applications.

Parkinson’s disease (PD) is a progressive neurodegenerative disorder characterized by bradykinesia, rigidity, and tremor. Considerable progress has been made to understand the exact mechanism leading to this disease. Most of what is known comes from the evidence of PD brains’ autopsies showing a deposition of Lewy bodies—containing a protein called α-synuclein (α-syn)—as the pathological determinant of PD. α-syn predisposes neurons to neurotoxicity and cell death, while the other associated mechanisms are mitochondrial dysfunction and oxidative stress, which are underlying precursors to the death of dopaminergic neurons at the substantia nigra pars compacta leading to disease progression. Several mechanisms have been proposed to unravel the pathological cascade of these diseases; most of them share a particular similarity: cell-to-cell communication through exosomes (EXOs). EXOs are intracellular membrane-based vesicles with diverse compositions involved in biological and pathological processes, which their secretion is driven by the NLR family pyrin domain-containing three proteins (NLRP3) inflammasome. Toxic biological fibrils are transferred to recipient cells, and the disposal of damaged organelles through generating mitochondrial-derived vesicles are suggested mechanisms for developing PD. EXOs carry various biomarkers; thus, they are promising to diagnose different neurological disorders, including neurodegenerative diseases (NDDs). As nanovesicles, the applications of EXOs are not only restricted as diagnostics but also expanded to treat NDDs as therapeutic carriers and nano-scavengers. Herein, the aim is to highlight the potential incrimination of EXOs in the pathological cascade and progression of PD and their role as biomarkers and therapeutic carriers for diagnosing and treating this neuro-debilitating disorder.

MIROs and DRP1 drive mitochondrial-derived vesicle biogenesis and promote quality control.

Mitochondrial-derived vesicles (MDVs) are implicated in diverse physiological processes-for example, mitochondrial quality control-and are linked to various neurodegenerative diseases. However, their specific cargo composition and complex molecular biogenesis are still unknown. Here we report the proteome and lipidome of steady-state TOMM20+ MDVs. We identified 107 high-confidence MDV cargoes, which include all β-barrel proteins and the TOM import complex. MDV cargoes are delivered as fully assembled complexes to lysosomes, thus representing a selective mitochondrial quality control mechanism for multi-subunit complexes, including the TOM machinery. Moreover, we define key biogenesis steps of phosphatidic acid-enriched MDVs starting with the MIRO1/2-dependent formation of thin membrane protrusions pulled along microtubule filaments, followed by MID49/MID51/MFF-dependent recruitment of the dynamin family GTPase DRP1 and finally DRP1-dependent scission. In summary, we define the function of MDVs in mitochondrial quality control and present a mechanistic model for global GTPase-driven MDV biogenesis.

Mitophagy: At the heart of mitochondrial quality control in cardiac aging and frailty.

Cardiovascular disease is highly prevalent among older adults and poses a huge burden on morbidity, disability, and mortality. The age-related increased vulnerability of the cardiovascular system towards stressors is a pathophysiological trait of cardiovascular disease. This has been associated with a progressive deterioration of blood vessels and decline in heart function during aging. Cardiomyocytes rely mostly on oxidative metabolism for deploying their activities and mitochondrial metabolism is crucial to this purpose. Dysmorphic, inefficient, and oxidant-producing mitochondria have been identified in aged cardiomyocytes in association with cardiac structural and functional alterations. These aberrant organelles are thought to arise from inefficient mitochondrial quality control, which has therefore been place in the spotlight as a relevant mechanism of cardiac aging. As a result of alterations in mitochondrial quality control and redox dyshomeostasis, mitochondrial damage accumulates and contributes to cardiac frailty. Herein, we discuss the contribution of defective mitochondrial quality control pathways to cardiac frailty. Emerging findings pointing towards the exploitation of these pathways as therapeutic targets against cardiac aging and cardiovascular disease will also be illustrated.

An online gathering about the latest on molecular membrane biology

An online gathering about the latest on molecular membrane biology

Chromatin remodeling due to degradation of citrate carrier impairs osteogenesis of aged mesenchymal stem cells

Aging is accompanied by a general decline in the function of many cellular pathways. However, whether these are causally or functionally interconnected remains elusive. Here, we study the effect of mitochondrial–nuclear communication on stem cell aging. We show that aged mesenchymal stem cells exhibit reduced chromatin accessibility and lower histone acetylation, particularly on promoters and enhancers of osteogenic genes. The reduced histone acetylation is due to impaired export of mitochondrial acetyl-CoA, owing to the lower levels of citrate carrier (CiC). We demonstrate that aged cells showed enhanced lysosomal degradation of CiC, which is mediated via mitochondrial-derived vesicles. Strikingly, restoring cytosolic acetyl-CoA levels either by exogenous CiC expression or via acetate supplementation, remodels the chromatin landscape and rescues the osteogenesis defects of aged mesenchymal stem cells. Collectively, our results establish a tight, age-dependent connection between mitochondrial quality control, chromatin and stem cell fate, which are linked together by CiC.

Extracellular Mitochondrial Components and Effects on Cardiovascular Disease.

Besides being powerhouses of the cell, mitochondria released into extracellular space act as intercellular signaling. Mitochondria and their components mediate cell-to-cell communication in free form or embedded in a carrier. The pathogenesis of cardiovascular disease is complex, which shows close relationship with inflammation and metabolic abnormalities. Since mitochondria sustain optimal function of the heart, extracellular mitochondria are emerging as a key regulator in the development of cardiovascular disease. In this review, we provide recent findings in the presence and forms of mitochondria transfer between cells, as well as the effects of these mitochondria on vascular inflammation and ischemic myocardium. Mitochondrial transplantation is a novel treatment paradigm for patients suffering from acute cardiovascular accident and challenges the traditional methods of mitochondria isolation.

Mitochondrial Dynamics and Mitophagy in Skeletal Muscle Health and Aging.

The maintenance of mitochondrial integrity is critical for muscle health. Mitochondria, indeed, play vital roles in a wide range of cellular processes, including energy supply, Ca2+ homeostasis, retrograde signaling, cell death, and many others. All mitochondria-containing cells, including skeletal muscle cells, dispose of several pathways to maintain mitochondrial health, including mitochondrial biogenesis, mitochondrial-derived vesicles, mitochondrial dynamics (fusion and fission process shaping mitochondrial morphology), and mitophagy—the process in charge of the removal of mitochondria though autophagy. The loss of skeletal muscle mass (atrophy) is a major health problem worldwide, especially in older people. Currently, there is no treatment to counteract the progressive decline in skeletal muscle mass and strength that occurs with aging, a process termed sarcopenia. There is increasing data, including our own, suggesting that accumulation of dysfunctional mitochondria contributes to the development of sarcopenia. Impairments in mitochondrial dynamics and mitophagy were recently proposed to contribute to sarcopenia. This review summarizes the current state of knowledge on the role played by mitochondrial dynamics and mitophagy in skeletal muscle health and in the development of sarcopenia. We also highlight recent studies showing that enhancing mitophagy in skeletal muscle is a promising therapeutic target to prevent or even treat skeletal muscle dysfunction in the elderly.

A new functional role of mitochondria-anchored protein ligase in peroxisome morphology in mammalian cells.

Mitochondria and peroxisomes are metabolically interconnected and functionally active subcellular organelles. These two dynamic organelles, share a number of common biochemical functions such as β-oxidation of fatty acids and detoxification of peroxides. The biogenesis and morphology of both these organelles in the mammalian cells is controlled by common transcription factors like PGC1α, and by a common fission machinery comprising of fission proteins like DRP1, Mff, and hFis1, respectively. In addition, the outer membrane mitochondria-anchored protein ligase (MAPL), the first mitochondrial SUMO E3 ligase with a RING-finger domain, also regulates mitochondrial morphology inducing mitochondrial fragmentation upon its overexpression. This fragmentation is dependent on both the RING domain of MAPL and the presence of the mitochondrial fission GTPase dynamin-related protein-1 (DRP1). Earlier studies have demonstrated that mitochondrial-derived vesiclesare formed independently of the known mitochondrial fission GTPase, DRP1 are enriched for MAPL and are targeted to peroxisomes. The current study shows that MAPL regulates morphology of peroxisomes in a cell-type specific manner. Fascinatingly, the peroxisome elongation caused either due to silencing of DRP1 or by addition of polyunsaturated fatty acid, docosahexaenoic acidwas blocked by overexpressing MAPL in mammalian cell lines. Furthermore, the transfection and colocalisation studies of MAPL with peroxisome membrane marker, PMP70, in different cell lines clearly revealed a cell-type specificity of transport of MAPL to peroxisomes. Previous work has placed the Vps35 (retromer component) as vital for delivery of MAPL to peroxisomes, placing the retromer as critical for the formation of MAPL-positive mitochondrial-derived vesicles. The results of polyethylene glycol-basedcell-cell fusion assay signified that the enrichment of MAPL in peroxisomes is through vesicles and a retromer dependent phenomenon. Thus, a novel function for MAPL in peroxisomes is established to regulate peroxisome elongation and morphology under growth conditions and thus possibly modulate peroxisome fission.

Mitochondrial-derived vesicles compensate for loss of LC3-mediated mitophagy.

Mitochondria are critical metabolic and signaling hubs, and dysregulated mitochondrial homeostasis is implicated in many diseases. Degradation of damaged mitochondria by selective GABARAP/LC3-dependent macro-autophagy (mitophagy) is critical for maintaining mitochondrial homeostasis. To identify alternate forms of mitochondrial quality control that functionally compensate if mitophagy is inactive, we selected for autophagy-dependent cancer cells that survived loss of LC3-dependent autophagosome formation caused by inactivation of ATG7 or RB1CC1/FIP200. We discovered rare surviving autophagy-deficient clones that adapted to maintain mitochondrial homeostasis after gene inactivation and identified two enhanced mechanisms affecting mitochondria including mitochondrial dynamics and mitochondrial-derived vesicles (MDVs). To further understand these mechanisms, we quantified MDVs via flow cytometry and confirmed an SNX9-mediated mechanism necessary for flux of MDVs to lysosomes. We show that the autophagy-dependent cells acquire unique dependencies on these processes, indicating that these alternate forms of mitochondrial homeostasis compensate for loss of autophagy to maintain mitochondrial health.

Proteomics characterization of mitochondrial-derived vesicles under oxidative stress.

Mitochondria share attributes of vesicular transport with their bacterial ancestors given their ability to form mitochondrial‐derived vesicles (MDVs). MDVs are involved in mitochondrial quality control and their formation is enhanced with stress and may, therefore, play a potential role in mitochondrial‐cellular communication. However, MDV proteomic cargo has remained mostly undefined. In this study, we strategically used an in vitro MDV budding/reconstitution assay on cardiac mitochondria, followed by graded oxidative stress, to identify and characterize the MDV proteome. Our results confirmed previously identified cardiac MDV markers, while also revealing a complete map of the MDV proteome, paving the way to a better understanding of the role of MDVs. The oxidative stress vulnerability of proteins directed the cargo loading of MDVs, which was enhanced by antimycin A (Ant‐A). Among OXPHOS complexes, complexes III and V were found to be Ant‐A‐sensitive. Proteins from metabolic pathways such as the TCA cycle and fatty acid metabolism, along with Fe‐S cluster, antioxidant response proteins, and autophagy were also found to be Ant‐A sensitive. Intriguingly, proteins containing hyper‐reactive cysteine residues, metabolic redox switches, including professional redox enzymes and those that mediate iron metabolism, were found to be components of MDV cargo with Ant‐A sensitivity. Last, we revealed a possible contribution of MDVs to the formation of extracellular vesicles, which may indicate mitochondrial stress. In conclusion, our study provides an MDV proteomics signature that delineates MDV cargo selectivity and hints at the potential for MDVs and their novel protein cargo to serve as vital biomarkers during mitochondrial stress and related pathologies.

The Role of Mitophagy in Regulating Cell Death.

Mitochondria are multifaceted organelles that serve to power critical cellular functions, including act as power generators of the cell, buffer cytosolic calcium overload, production of reactive oxygen species, and modulating cell survival. The structure and the cellular location of mitochondria are critical for their function and depend on highly regulated activities such as mitochondrial quality control (MQC) mechanisms. The MQC is regulated by several sets of processes: mitochondrial biogenesis, mitochondrial fusion and fission, mitophagy, and other mitochondrial proteostasis mechanisms such as mitochondrial unfolded protein response (mtUPR) or mitochondrial-derived vesicles (MDVs). These processes are important for the maintenance of mitochondrial homeostasis, and alterations in the mitochondrial function and signaling are known to contribute to the dysregulation of cell death pathways. Recent studies have uncovered regulatory mechanisms that control the activity of the key components for mitophagy. In this review, we discuss how mitophagy is controlled and how mitophagy impinges on health and disease through regulating cell death.

A central role for mitochondrial-derived vesicles in the innate immune response: implications for Parkinson's disease.

A central role for mitochondrial-derived vesicles in the innate immune response: implications for Parkinson's disease.

Proteomic Profiling of Mitochondrial-Derived Vesicles in Brain Reveals Enrichment of Respiratory Complex Sub-assemblies and Small TIM Chaperones.

The generation of mitochondrial-derived vesicles (MDVs) is implicated in a plethora of vital cell functions, from mitochondrial quality control to peroxisomal biogenesis. The discovery of distinct subtypes of MDVs has revealed the selective inclusion of mitochondrial cargo in response to varying stimuli. However, the true scope and variety of MDVs is currently unclear, and unbiased approaches have yet to be used to understand their biology. Furthermore, as mitochondrial dysfunction has been implicated in many neurodegenerative diseases, it is essential to understand MDV pathways in the nervous system. To address this, we sought to identify the cargo in brain MDVs. We used an in vitro budding assay and proteomic approach to identify proteins selectively enriched in MDVs. 72 proteins were identified as MDV-enriched, of which 31% were OXPHOS proteins. Interestingly, the OXPHOS proteins localized to specific modules of the respiratory complexes, hinting at the inclusion of sub-assemblies in MDVs. Small TIM chaperones were also highly enriched in MDVs, linking mitochondrial chaperone-mediated protein transport to MDV formation. As the two Parkinson's disease genes PINK1 and Parkin have been previously implicated in MDV biogenesis in response to oxidative stress, we compared the MDV proteomes from the brains of wild-type mice with those of PINK1-/- and Parkin-/- mice. No significant difference was found, suggesting that PINK1- and Parkin-dependent MDVs make up a small proportion of all MDVs in the brain. Our findings demonstrate a previously uncovered landscape of MDV complexity and provide a foundation from which further novel MDV functions can be discovered. Data are available via ProteomeXchange with identifier PXD020197.

Extracellular Vesicles and Damage-Associated Molecular Patterns: A Pandora's Box in Health and Disease.

Sterile inflammation develops as part of an innate immunity response to molecules released upon tissue injury and collectively indicated as damage-associated molecular patterns (DAMPs). While coordinating the clearance of potential harmful stimuli, promotion of tissue repair, and restoration of tissue homeostasis, a hyper-activation of such an inflammatory response may be detrimental. The complex regulatory pathways modulating DAMPs generation and trafficking are actively investigated for their potential to provide relevant insights into physiological and pathological conditions. Abnormal circulating extracellular vesicles (EVs) stemming from altered endosomal-lysosomal system have also been reported in several age-related conditions, including cancer and neurodegeneration, and indicated as a promising route for therapeutic purposes. Along this pathway, mitochondria may dispose altered components to preserve organelle homeostasis. However, whether a common thread exists between DAMPs and EVs generation is yet to be clarified. A deeper understanding of the highly complex, dynamic, and variable intracellular and extracellular trafficking of DAMPs and EVs, including those of mitochondrial origin, is needed to unveil relevant pathogenic pathways and novel targets for drug development. Herein, we describe the mechanisms of generation of EVs and mitochondrial-derived vesicles along the endocytic pathway and discuss the involvement of the endosomal-lysosomal in cancer and neurodegeneration (i.e., Alzheimer’s and Parkinson’s disease).

Regulation of Parkin-dependent mitophagy by Bcl-2-associated athanogene (BAG) family members.

Regulation of Parkin-dependent mitophagy by Bcl-2-associated athanogene (BAG) family members.

Circulating Mitochondrial-Derived Vesicles, Inflammatory Biomarkers and Amino Acids in Older Adults With Physical Frailty and Sarcopenia: A Preliminary BIOSPHERE Multi-Marker Study Using Sequential and Orthogonalized Covariance Selection - Linear Discriminant Analysis.

Physical frailty and sarcopenia (PF&S) is a prototypical geriatric condition characterized by reduced physical function and low muscle mass. The multifaceted pathophysiology of this condition recapitulates all hallmarks of aging making the identification of specific biomarkers challenging. In the present study, we explored the relationship among three processes that are thought to be involved in PF&S (i.e., systemic inflammation, amino acid dysmetabolism, and mitochondrial dysfunction). We took advantage of the well-characterized cohort of older adults recruited in the “BIOmarkers associated with Sarcopenia and Physical frailty in EldeRly pErsons” (BIOSPHERE) study to preliminarily combine in a multi-platform analytical approach inflammatory biomolecules, amino acids and derivatives, and mitochondrial-derived vesicle (MDV) cargo molecules to evaluate their performance as possible biomarkers for PF&S. Eleven older adults aged 70 years and older with PF&S and 10 non-sarcopenic non-frail controls were included in the analysis based on the availability of the three categories of biomolecules. A sequential and orthogonalized covariance selection—linear discriminant analysis (SO-CovSel–LDA) approach was used for biomarkers selection. Of the 75 analytes assayed, 16 had concentrations below the detection limit. Within the remaining 59 biomolecules, So-CovSel–LDA selected a set comprising two amino acids (phosphoethanolamine and tryptophan), two cytokines (interleukin 1 receptor antagonist and macrophage inflammatory protein 1β), and MDV-derived nicotinamide adenine dinucleotide reduced form:ubiquinone oxidoreductase subunit S3 as the best predictors for discriminating older people with and without PF&S. The evaluation of these biomarkers in larger cohorts and their changes over time or in response to interventions may unveil specific pathogenetic pathways of PF&S and identify new biological targets for drug development.