Mitochondrial permeability transition (mPT) is a phenomenon that abruptly causes the flux of low molecular weight solutes (molecular weight up to 1,500) across the generally impermeable inner ...mitochondrial membrane. The mPT is mediated by the so-called mitochondrial permeability transition pore (mPTP), a supramolecular entity assembled at the interface of the inner and outer mitochondrial membranes. In contrast to mitochondrial outer membrane permeabilization, which mostly activates apoptosis, mPT can trigger different cellular responses, from the physiological regulation of mitophagy to the activation of apoptosis or necrosis. Although there are several molecular candidates for the mPTP, its molecular nature remains contentious. This lack of molecular data was a significant setback that prevented mechanistic insight into the mPTP, pharmacological targeting and the generation of informative animal models. In recent years, experimental evidence has highlighted mitochondrial F
F
ATP synthase as a participant in mPTP formation, although a molecular model for its transition to the mPTP is still lacking. Recently, the resolution of the F
F
ATP synthase structure by cryogenic electron microscopy led to a model for mPTP gating. The elusive molecular nature of the mPTP is now being clarified, marking a turning point for understanding mitochondrial biology and its pathophysiological ramifications. This Review provides an up-to-date reference for the understanding of the mammalian mPTP and its cellular functions. We review current insights into the molecular mechanisms of mPT and validated observations - from studies in vivo or in artificial membranes - on mPTP activity and functions. We end with a discussion of the contribution of the mPTP to human disease. Throughout the Review, we highlight the multiple unanswered questions and, when applicable, we also provide alternative interpretations of the recent discoveries.
A large body of evidence indicates that mitochondrial dysfunction has a major role in the pathogenesis of multiple cardiovascular disorders. Over the past 2 decades, extraordinary efforts have been ...focused on the development of agents that specifically target mitochondria for the treatment of cardiovascular disease. Despite such an intensive wave of investigation, no drugs specifically conceived to modulate mitochondrial functions are currently available for the clinical management of cardiovascular disease. In this Review, we discuss the therapeutic potential of targeting mitochondria in patients with cardiovascular disease, examine the obstacles that have restrained the development of mitochondria-targeting agents thus far, and identify strategies that might empower the full clinical potential of this approach.
Membrane contact sites between virtually any known organelle have been documented and, in the last decades, their study received momentum due to their importance for fundamental activities of the ...cell and for the subtle comprehension of many human diseases. The lack of tools to finely image inter-organelle proximity hindered our understanding on how these subcellular communication hubs mediate and regulate cell homeostasis. We develop an improved and expanded palette of split-GFP-based contact site sensors (SPLICS) for the detection of single and multiple organelle contact sites within a scalable distance range. We demonstrate their flexibility under physiological conditions and in living organisms.
Since its discovery in the 1970s, the mitochondrial permeability transition (MPT) has been proposed to be a strategic regulator of cell death. Intense research efforts have focused on elucidating the ...molecular components of the MPT because this knowledge may help to better understand and treat various pathologies ranging from neurodegenerative and cardiac diseases to cancer. In the case of cancer, several studies have revealed alterations in the activity of the mitochondrial permeability transition pore (mPTP) and have determined its regulatory mechanism; these studies have also suggested that suppression of the activity of the mPTP, rather than its inactivation, commonly occurs in solid neoplasms. This review focuses on the most recent advances in understanding mPTP regulation in cancer and highlights the ability of the mPTP to impede the mechanisms of cell death.
Mitochondrial permeability transition (mPT) refers to a sudden increase in the permeability of the inner mitochondrial membrane. Long-term studies of mPT revealed that this phenomenon has a critical ...role in multiple pathophysiological processes. mPT is mediated by the opening of a complex termed the mPT pore (mPTP), which is responsible for the osmotic influx of water into the mitochondrial matrix, resulting in swelling of mitochondria and dissipation of the mitochondrial membrane potential. Here we provide three independent optimized protocols for monitoring mPT in living cells: (i) measurement using a calcein-cobalt technique, (ii) measurement of the mPTP-dependent alteration of the mitochondrial membrane potential, and (iii) measurement of mitochondrial swelling. These procedures can easily be modified and adapted to different cell types. Cell culture and preparation of the samples are estimated to take ∼1 d for methods (i) and (ii), and ∼3 d for method (iii). The entire experiment, including analyses, takes ∼2 h.
Mitochondria are considered one of the main sources of reactive oxygen species (ROS). The overgeneration of ROS can evoke an intracellular state of oxidative stress, leading to permanent cell damage. ...Thus, the intracellular accumulation of ROS may not only disrupt the functions of specific tissues and organs but also lead to the premature death of the entire organism. Less severe increases in ROS levels may lead to the nonlethal oxidation of fundamental cellular components, such as proteins, phospholipids, and DNA, hence exerting a mutagenic effect that promotes oncogenesis and tumor progression. Here, we describe the use of chemical probes for the rapid detection of ROS in intact and permeabilized adherent cells by fluorescence microscopy and fluorometry. Moreover, after discussing the limitations described in the literature for the fluorescent probes presented herein, we recommend methods to assess the production of specific ROS in various fields of investigation, including the study of oncometabolism.
Abstract The mitochondrial permeability transition is a key event in cell death. Intense research efforts have been focused on elucidating the molecular components of the mitochondrial permeability ...transition pore (mPTP) to improve the understanding and treatment of various pathologies, including neurodegenerative disorders, cancer and cardiac diseases. Several molecular factors have been proposed as core components of the mPTP; however, further investigation has indicated that these factors are among a wide range of regulators. Thus, the scientific community lacks a clear model of the mPTP. Here, we review the molecular factors involved in the regulation and formation of the mPTP. Furthermore, we propose that the mitochondrial ATP synthase, specifically its c subunit, is the central core component of the mPTP complex. Moreover, we discuss the involvement of the mPTP in ischemia and reperfusion as well as the results of clinical studies targeting the mPTP to ameliorate ischemia-reperfusion injury. This article is part of a Special Issue entitled "Mitochondria: From Basic Mitochondrial Biology to Cardiovascular Disease".
The impact of the mitochondrial permeability transition (MPT) on cellular physiology is well characterized. In contrast, the composition and mode of action of the permeability transition pore complex ...(PTPC), the supramolecular entity that initiates MPT, remain to be elucidated. Specifically, the precise contribution of the mitochondrial F1FO ATP synthase (or subunits thereof) to MPT is a matter of debate. We demonstrate that F1FO ATP synthase dimers dissociate as the PTPC opens upon MPT induction. Stabilizing F1FO ATP synthase dimers by genetic approaches inhibits PTPC opening and MPT. Specific mutations in the F1FO ATP synthase c subunit that alter C‐ring conformation sensitize cells to MPT induction, which can be reverted by stabilizing F1FO ATP synthase dimers. Destabilizing F1FO ATP synthase dimers fails to trigger PTPC opening in the presence of mutants of the c subunit that inhibit MPT. The current study does not provide direct evidence that the C‐ring is the long‐sought pore‐forming subunit of the PTPC, but reveals that PTPC opening requires the dissociation of F1FO ATP synthase dimers and involves the C‐ring.
Synopsis
F1FO ATP synthase dimer dissociation plays a crucial role in MPT and allows PTPC opening. In the same context, c subunit‐specific mutations altering the C‐ring conformation prime cells for MPT induction.
MPT is linked with dissociation of F1FO ATP synthase dimers and their stabilization by ATPIF1 or ATP5I overexpression inhibits PTPC opening.
The glycine zipper domain of the c subunit plays a central role in the MPT process as specific mutations promoting conformational changes in the C‐ring structure affect PTPC opening.
A proper C‐ring conformation is required for MPT induction once F1FO ATP synthase dimers have dissociated.
F1FO ATP synthase dimer dissociation plays a crucial role in MPT and allows PTPC opening. In the same context, c subunit‐specific mutations altering the C‐ring conformation prime cells for MPT induction.
Mitochondria are considered as the main source of reactive oxygen species (ROS) in the cell. For this reason, they have been recognized as a source of various pathological conditions as well as ...aging. Chronic increase in the rate of ROS production is responsible for the accumulation of ROS-associated damages in DNA, proteins, and lipids, and may result in progressive cell dysfunctions and, in a consequence, apoptosis, increasing the overall probability of an organism’s pathological conditions. The superoxide anion is the main undesired by-product of mitochondrial oxidative phosphorylation. Its production is triggered by a leak of electrons from the mitochondrial respiratory chain and the reaction of these electrons with O2. Superoxide dismutase (MnSOD, SOD2) from the mitochondrial matrix as well as superoxide dismutase (Cu/ZnSOD, SOD1) present in small amounts in the mitochondrial intramembrane space, convert superoxide anion to hydrogen peroxide, which can be then converted by catalase to harmless H2O. In this chapter, we describe a relation between mitochondrial membrane potential and the rate of ROS formation. We present different methods applicable for isolated mitochondria or intact cells. We also present experiments demonstrating that a magnitude and a direction (increase or decrease) of a change in mitochondrial ROS production depends on the metabolic state of this organelle.
Mitochondria are considered the main source of reactive oxygen species (ROS) in the cell. For this reason they have been recognized as a source of various pathological conditions as well as aging. ...Chronic increase in the rate of ROS production is responsible for the accumulation of ROS-associated damages in DNA, proteins, and lipids and may result in progressive cell dysfunctions and, in a consequence, apoptosis, increasing the overall probability of an organism's pathological conditions. The superoxide anion is the main undesired by-product of mitochondrial oxidative phosphorylation. Its production is triggered by a leak of electrons from the mitochondrial respiratory chain and the reaction of these electrons with O
. Superoxide dismutase (MnSOD, SOD2) from the mitochondrial matrix, as well as superoxide dismutase (Cu/ZnSOD, SOD1) present in small amounts in the mitochondrial intramembrane space, converts superoxide anion to hydrogen peroxide, which can be then converted by catalase to harmless H
O.In the chapter we describe a relation between mitochondrial membrane potential and the rate of ROS formation. We present different methods applicable for isolated mitochondria or intact cells. We also present experiments demonstrating that a magnitude and a direction (increase or decrease) of a change in mitochondrial ROS production depend on the metabolic state of this organelle.
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