Mechanisms Orchestrating Mitochondrial Dynamics for Energy Homeostasis

To maintain homeostasis, every cell must constantly monitor its energy level and appropriately adjust energy, in the form of ATP, production rates based on metabolic demand. Continuous fulfillment of this energy demand depends on the ability of cells to sense, metabolize, and convert nutrients into chemical energy. Mitochondria are the main energy conversion sites for many cell types. Cellular metabolic states dictate the mitochondrial size, shape, function, and positioning. Mitochondrial shape varies from singular discrete organelles to interconnected reticular networks within cells. The morphological adaptations of mitochondria to metabolic cues are facilitated by the dynamic events categorized as transport, fusion, fission, and quality control. By changing their dynamics and strategic positioning within the cytoplasm, mitochondria carry out critical functions and also participate actively in inter-organelle cross-talk, assisting metabolite transfer, degradation, and biogenesis. Mitochondrial dynamics has become an active area of research because of its particular importance in cancer, metabolic diseases, and neurological disorders. In this review, we will highlight the molecular pathways involved in the regulation of mitochondrial dynamics and their roles in maintaining energy homeostasis.     

Mitochondrial Ca(2+) and neurodegeneration

Mitochondria are essential for ensuring numerous fundamental physiological processes such as cellular energy, redox balance, modulation of Ca(2+) signaling and important biosynthetic pathways. They also govern the cell fate by participating in the apoptosis pathway. The mitochondrial shape, volume, number and distribution within the cells are strictly controlled. The regulation of these parameters has an impact on mitochondrial function, especially in the central nervous system, where trafficking of mitochondria is critical to their strategic intracellular distribution, presumably according to local energy demands. Thus, the maintenance of a healthy mitochondrial population is essential to avoid the impairment of the processes they regulate: for this purpose, cells have developed mechanisms involving a complex system of quality control to remove damaged mitochondria, or to renew them. Defects of these processes impair mitochondrial function and lead to disordered cell function, i.e., to a disease condition. Given the standard role of mitochondria in all cells, it might be expected that their dysfunction would give rise to similar defects in all tissues. However, damaged mitochondrial function has pleiotropic effects in multicellular organisms, resulting in diverse pathological conditions, ranging from cardiac and brain ischemia, to skeletal muscle myopathies to neurodegenerative diseases. In this review, we will focus on the relationship between mitochondrial (and cellular) derangements and Ca(2+) dysregulation in neurodegenerative diseases, emphasizing the evidence obtained in genetic models. Common patterns, that recognize the derangement of Ca(2+) and energy control as a causative factor, have been identified: advances in the understanding of the molecular regulation of Ca(2+) homeostasis, and on the ways in which it could become perturbed in neurological disorders, may lead to the development of therapeutic strategies that modulate neuronal Ca(2+) signaling.     

Mitochondrial respiratory chain dysfunction: implications in neurodegeneration

For decades mitochondria have been considered static round-shaped organelles in charge of energy production. In contrast, they are highly dynamic cellular components that undergo continuous cycles of fusion and fission influenced, for instance, by oxidative stress, cellular energy requirements, or the cell cycle state. New important functions beyond energy production have been attributed to mitochondria, such as the regulation of cell survival, because of their role in the modulation of apoptosis, autophagy, and aging. Primary mitochondrial diseases due to mutations in genes involved in these new mitochondrial functions and the implication of mitochondrial dysfunction in multifactorial human pathologies such as cancer, Alzheimer and Parkinson diseases, or diabetes has been demonstrated. Therefore, mitochondria are set at a central point of the equilibrium between health and disease, and a better understanding of mitochondrial functions will open new fields for exploring the roles of these mitochondrial pathways in human pathologies. This review dissects the relationships between activity and assembly defects of the mitochondrial respiratory chain, oxidative damage, and alterations in mitochondrial dynamics, with special focus on their implications for neurodegeneration.     

Mitochondrial DNA (mtDNA) Biogenesis: Visualization and Duel Incorporation of BrdU and EdU Into Newly Synthesized mtDNA In Vitro

Mitochondria are key regulators of cellular energy and are the focus of a large number of studies examining the regulation of mitochondrial dynamics and biogenesis in healthy and diseased conditions. One approach to monitoring mitochondrial biogenesis is to measure the rate of mitochondrial DNA (mtDNA) replication. We developed a sensitive technique to visualize newly synthesized mtDNA in individual cells to study mtDNA replication within subcellular compartments of neurons. The technique combines the incorporation of 5-bromo-2-deoxyuridine (BrdU) and/or 5-ethynyl-2′-deoxyuridine (EdU) into mtDNA, together with a tyramide signal amplification protocol. Employing this technique, we visualized and measured mtDNA biogenesis in individual cells. The labeling procedure for EdU allows for more comprehensive results by allowing the comparison of its incorporation with other intracellular markers, because it does not require the harsh acid or enzyme digests necessary to recover the BrdU epitope. In addition, the utilization of both BrdU and EdU permits sequential pulse–chase experiments to follow the intracellular localization of mtDNA replication. The ability to quantify mitochondrial biogenesis provides an essential tool for investigating the alterations in mitochondrial dynamics involved in the pathogenesis of multiple cellular disorders, including neuropathies and neurodegenerative diseases. (J Histochem Cytochem 58:207–218, 2010)     

PGC-1α的转录调节和翻译后修饰

过氧化物酶体增殖物激活受体γ(peroxisome proliferator activated receptorγ,PPARγ)辅助激活因子-1α(PPARγcoactivator-1α,PGC-1α)是线粒体生物合成的关键调节分子.外界刺激(寒冷、饥饿、运动)一方面可以改变PGC-1α的基因和蛋白质表达水平,另一方面可以通过翻译后修饰方式调节其蛋白质活性,最终调节细胞能量代谢和线粒体生物合成过程.PGC-1α表达的异常是代谢性疾病及老年性疾病等发病的重要原因.本文就PGC-1α在转录水平和翻译后修饰水平的调节方式的最新研究进展作一综述.     

Monitoring mitochondrial translation in living cells

Mitochondria possess a small genome that codes for core subunits of the oxidative phosphorylation system and whose expression is essential for energy production. Information on the regulation and spatial organization of mitochondrial gene expression in the cellular context has been difficult to obtain. Here we devise an imaging approach to analyze mitochondrial translation within the context of single cells, by following the incorporation of clickable non‐canonical amino acids. We apply this method to multiple cell types, including specialized cells such as cardiomyocytes and neurons, and monitor with spatial resolution mitochondrial translation in axons and dendrites. We also show that translation imaging allows to monitor mitochondrial protein expression in patient fibroblasts. Approaching mitochondrial translation with click chemistry opens new avenues to understand how mitochondrial biogenesis is integrated into the cellular context and can be used to assess mitochondrial gene expression in mitochondrial diseases.     

Control of mitochondrial integrity in ageing and disease

Various molecular and cellular pathways are active in eukaryotes to control the quality and integrity of mitochondria. These pathways are involved in keeping a ‘healthy’ population of this essential organelle during the lifetime of the organism. Quality control (QC) systems counteract processes that lead to organellar dysfunction manifesting as degenerative diseases and ageing. We discuss disease- and ageing-related pathways involved in mitochondrial QC: mtDNA repair and reorganization, regeneration of oxidized amino acids, refolding and degradation of severely damaged proteins, degradation of whole mitochondria by mitophagy and finally programmed cell death. The control of the integrity of mtDNA and regulation of its expression is essential to remodel single proteins as well as mitochondrial complexes that determine mitochondrial functions. The redundancy of components, such as proteases, and the hierarchies of the QC raise questions about crosstalk between systems and their precise regulation. The understanding of the underlying mechanisms on the genomic, proteomic, organellar and cellular levels holds the key for the development of interventions for mitochondrial dysfunctions, degenerative processes, ageing and age-related diseases resulting from impairments of mitochondria.     

Parkin介导的线粒体自噬及细胞器互作机制

线粒体是细胞生理代谢活动发生的重要场所. 线粒体生发降解平衡是维持能量代谢稳定的重要保障. Parkin作为E3泛素连接酶，通过PINK1/Parkin、LC3等多种信号参与调控线粒体自噬过程. 此外，Parkin还能够影响线粒体相关内质网膜、调控细胞器间钙流，在线粒体-内质网对话过程中调控溶酶体途径介导的线粒体自噬. 脂肪组织是研究线粒体调节机制的理想模型：寒冷刺激诱导富含线粒体的米色脂肪生成；移除刺激后，组织中线粒体消失恢复为白色脂肪，但线粒体稳定性的调控机理目前仍有很多未知. 本文综述Parkin介导线粒体自噬途径的最新研究进展，及其参与线粒体、内质网、溶酶体等不同细胞器间相互作用的调控机制.     

Ubiquitin-dependent mitochondrial protein degradation

Progressive mitochondrial failure is tightly associated with the onset of many age-related human pathologies. This tight connection results from the double-edged sword of mitochondrial respiration, which is responsible for generating both ATP and ROS, as well as from risks that are inherent to mitochondrial biogenesis. To prevent and treat these diseases, a precise understanding of the mechanisms that maintain functional mitochondria is necessary. Mitochondrial protein quality control is one of the mechanisms that protect mitochondrial integrity, and increasing evidence implicates the cytosolic ubiquitin/proteasome system (UPS) as part of this surveillance network. In this review, we will discuss our current understanding of UPS-dependent mitochondrial protein degradation, its roles in diseases progression, and insights into future studies.     

Visualization of Mitochondrial DNA Replication in Individual Cells by EdU Signal Amplification

Mitochondria are key regulators of cellular energy and mitochondrial biogenesis is an essential component of regulating mitochondria numbers in healthy cells^1-3. One approach for monitoring mitochondrial biogenesis is to measure the rate of mitochondrial DNA (mtDNA) replication⁴. We developed a sensitive technique to label newly synthesized mtDNA in individual cells in order to study mtDNA biogenesis. The technique combines the incorporation of 5-ethynyl-2''-deoxyuridine (EdU)^5-7 with a tyramide signal amplification (TSA)⁸ protocol to visualize mtDNA replication within subcellular compartments of neurons. EdU is superior to other thymidine analogs, such as 5-bromo-2-deoxyuridine (BrdU), because the initial click reaction to label EdU^5-7 does not require the harsh acid treatments or enzyme digests that are required for exposing the BrdU epitope. The milder labeling of EdU allows for direct comparison of its incorporation with other cellular markers^9-10. The ability to visualize and quantify mtDNA biogenesis provides an essential tool for investigating the mechanisms used to regulate mitochondrial biogenesis and would provide insight into the pathogenesis associated with drug toxicity, aging, cancer and neurodegenerative diseases. Our technique is applicable to sensory neurons as well as other cell types. The use of this technique to measure mtDNA biogenesis has significant implications in furthering the understanding of both normal cellular physiology as well as impaired disease states.     

Number matters: control of mammalian mitochondrial DNA copy number

Regulation of mitochondrial biogenesis is essential for proper cellular functioning. Mitochondrial DNA （mtDNA） depletion and the resulting mitochondrial malfunction have been implicated in cancer, neurodegeneration, diabetes, aging, and many other human diseases. Although it is known that the dynamics of the mammalian mitochondrial genome are not linked with that of the nuclear genome, very little is known about the mechanism of mtDNA propagation. Nevertheless, our understanding of the mode of mtDNA replication has ad- vanced in recent years, though not without some controversies. This review summarizes our current knowledge of mtDNA copy number control in mammalian cells, while focusing on both mtDNA replication and turnover. Although mtDNA copy number is seemingly in excess, we reason that mtDNA copy number control is an important aspect of mitochondrial genetics and biogenesis and is essential for normal cellular function.     

Two MAPK-signaling pathways are required for mitophagy in Saccharomyces cerevisiae

Macroautophagy (hereafter referred to simply as autophagy) is a catabolic pathway that mediates the degradation of long-lived proteins and organelles in eukaryotic cells. The regulation of mitochondrial degradation through autophagy plays an essential role in the maintenance and quality control of this organelle. Compared with our understanding of the essential function of mitochondria in many aspects of cellular metabolism such as energy production and of the role of dysfunctional mitochondria in cell death, little is known regarding their degradation and especially how upstream signaling pathways control this process. Here, we report that two mitogen-activated protein kinases (MAPKs), Slt2 and Hog1, are required for mitophagy in Saccharomyces cerevisiae. Slt2 is required for the degradation of both mitochondria and peroxisomes (via pexophagy), whereas Hog1 functions specifically in mitophagy. Slt2 also affects the recruitment of mitochondria to the phagophore assembly site (PAS), a critical step in the packaging of cargo for selective degradation.     

Mitochondrial quality control as a key determinant of cell survival

Mitochondria are the energy producing dynamic double-membraned organelles essential for cellular and organismal survival. A multitude of intra- and extra-cellular signals involved in the regulation of energy metabolism and cell fate determination converge on mitochondria to promote or prevent cell survival by modulating mitochondrial function and structure. Mitochondrial fitness is maintained by mitophagy, a pathway of selective degradation of dysfunctional organelles. Mitophagy impairment and altered clearance results in increased levels of dysfunctional and structurally aberrant mitochondria, changes in energy production, loss of responsiveness to intra- and extra-cellular signals and ultimately cell death. The decline of mitochondrial function and homeostasis with age is reported to be central to age-related pathologies. Here we discuss the molecular mechanisms controlling mitochondrial dynamics, mitophagy and cell death signalling and how their perturbation may contribute to ageing and age-related illness.