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Primary ovarian insufficiency (POI) is a rare gynecological condition. This disease causes menstrual disturbances, infertility, and various health problems. Historically, hormone replacement therapy is the first-line treatment for this disorder. Women diagnosed with POI are left with limited therapeutic options. In order to remedy this situation, a new generation of therapeutic approaches, such as in vitro activation, mitochondrial activation technique, stem cell and exosomes therapy, biomaterials strategies, and platelet-rich plasma intra-ovarian infusion, is being developed. However, these emerging therapies are yet in the experimental stage and require precise design components to accelerate their conversion into clinical treatments. Thus, each medical practitioner bears responsibility for selecting suitable therapies for individual patients. In this article, we provide a timely analysis of the therapeutic strategies that are available for POI patients and discuss the prospects of POI therapy.

Mitochondrial DNA (mtDNA) is a potent damage-associated molecular pattern that, if it reaches the cytoplasm or extracellular milieu, triggers innate immune pathways. mtDNA signaling has been implicated in a wide range of diseases; however, the mechanisms of mtDNA release are unclear, and the process has not been observed in real time thus far. McArthur et al. used live-cell lattice light-sheet microscopy to look at mtDNA release during intrinsic apoptosis. Activation of the pro-death proteins BAK and BAX resulted in the formation of large macro-pores in the mitochondrial outer membrane. These massive holes caused the inner mitochondrial membrane to balloon out into the cytoplasm, resulting in mitochondrial herniation. This process allowed the contents of the mitochondrial matrix, including mtDNA, to escape into the cytoplasm. Science , this issue p. eaao6047 Mitochondrial DNA is released from mitochondria in apoptotic cells as a result of BAK/BAX-induced mitochondrial herniation. Mitochondrial apoptosis is mediated by BAK and BAX, two proteins that induce mitochondrial outer membrane permeabilization, leading to cytochrome c release and activation of apoptotic caspases. In the absence of active caspases, mitochondrial DNA (mtDNA) triggers the innate immune cGAS/STING pathway, causing dying cells to secrete type I interferon. How cGAS gains access to mtDNA remains unclear. We used live-cell lattice light-sheet microscopy to examine the mitochondrial network in mouse embryonic fibroblasts. We found that after BAK/BAX activation and cytochrome c loss, the mitochondrial network broke down and large BAK/BAX pores appeared in the outer membrane. These BAK/BAX macropores allowed the inner mitochondrial membrane to herniate into the cytosol, carrying with it mitochondrial matrix components, including the mitochondrial genome. Apoptotic caspases did not prevent herniation but dismantled the dying cell to suppress mtDNA-induced innate immune signaling.

We found that a subset of signal transducer and activator of transcription 3 (STAT3) translocated into mitochondria in phagocytes, including macrophages isolated from individuals with sepsis. However, the role of mitochondrial STAT3 in macrophages remains unclear. To investigate the function of mitochondrial STAT3 , we generated inducible mitochondrial STAT3 knock-in mice. A cytokine array analysis, a CBA analysis, flow cytometry, immunofluorescence staining and quantification and metabolic analyses were subsequently performed in an LPS-induced sepsis model. Single-cell RNA sequencing, a microarray analysis, metabolic assays, mass spectrometry and ChIP assays were utilized to gain insight into the mechanisms of mitochondrial STAT3 in metabolic reprogramming in LPS-induced sepsis. We found that mitochondrial STAT3 induced NF-κB nuclear localization and exacerbated LPS-induced sepsis in parallel with a metabolic switch from mainly using glucose to an increased reliance on fatty acid oxidation (FAO). Moreover, mitochondrial STAT3 abrogated carnitine palmitoyl transferase 1a (CPT1a) ubiquitination and degradation in LPS-treated macrophages. Meanwhile, an interaction between CPT1a and ubiquitin-specific peptidase 50 (USP50) was observed. In contrast, knocking down USP50 decreased CPT1a expression and FAO mediated by mitochondrial STAT3. The ChIP assays revealed that NF-κB bound the USP50 promoter. Curcumin alleviated LPS-mediated sepsis by suppressing the activities of mitochondrial STAT3 and NF-κB. Our findings reveal that mitochondrial STAT3 could trigger FAO by inducing CPT1a stabilization mediated by USP50 in macrophages, at least partially.

Ageing in the worm Caenorhabditis elegans is shown to be delayed by supplementation with α-ketoglutarate, an effect that is probably mediated by ATP synthase—which is identified as a direct target of α-ketoglutarate—and target of rapamycin (TOR). How a small molecule extends C. elegans lifespan Calorie restriction can extend lifespan and delay age-related deterioration in a range of organisms. A few small-molecule metabolites have been shown to regulate the ageing process, but little is known about the mechanisms involved. Here Jing Huang and colleagues report that the tricarboxylic acid cycle intermediate α-ketoglutarate (α-KG) extends the lifespan of adult Caenorhabditis elegans roundworms by approximately 50%. The molecular target of α-KG is the β subunit of ATPase. α-KG is dependent on the TOR (target of rapamycin) pathway and it does not extend the lifespan of dietary-restricted animals, suggesting a link between the effects of α-KG and starvation/dietary restriction. Metabolism and ageing are intimately linked. Compared with ad libitum feeding, dietary restriction consistently extends lifespan and delays age-related diseases in evolutionarily diverse organisms 1 , 2 . Similar conditions of nutrient limitation and genetic or pharmacological perturbations of nutrient or energy metabolism also have longevity benefits 3 , 4 . Recently, several metabolites have been identified that modulate ageing 5 , 6 ; however, the molecular mechanisms underlying this are largely undefined. Here we show that α-ketoglutarate (α-KG), a tricarboxylic acid cycle intermediate, extends the lifespan of adult Caenorhabditis elegans . ATP synthase subunit β is identified as a novel binding protein of α-KG using a small-molecule target identification strategy termed drug affinity responsive target stability (DARTS) 7 . The ATP synthase, also known as complex V of the mitochondrial electron transport chain, is the main cellular energy-generating machinery and is highly conserved throughout evolution 8 , 9 . Although complete loss of mitochondrial function is detrimental, partial suppression of the electron transport chain has been shown to extend C. elegans lifespan 10 , 11 , 12 , 13 . We show that α-KG inhibits ATP synthase and, similar to ATP synthase knockdown, inhibition by α-KG leads to reduced ATP content, decreased oxygen consumption, and increased autophagy in both C. elegans and mammalian cells. We provide evidence that the lifespan increase by α-KG requires ATP synthase subunit β and is dependent on target of rapamycin (TOR) downstream. Endogenous α-KG levels are increased on starvation and α-KG does not extend the lifespan of dietary-restricted animals, indicating that α-KG is a key metabolite that mediates longevity by dietary restriction. Our analyses uncover new molecular links between a common metabolite, a universal cellular energy generator and dietary restriction in the regulation of organismal lifespan, thus suggesting new strategies for the prevention and treatment of ageing and age-related diseases.

Somatic mutations of ASXL1 are frequently detected in age-related clonal hematopoiesis (CH). However, how ASXL1 mutations drive CH remains elusive. Using knockin (KI) mice expressing a C-terminally truncated form of ASXL1-mutant (ASXL1-MT), we examined the influence of ASXL1-MT on physiological aging in hematopoietic stem cells (HSCs). HSCs expressing ASXL1-MT display competitive disadvantage after transplantation. Nevertheless, in genetic mosaic mouse model, they acquire clonal advantage during aging, recapitulating CH in humans. Mechanistically, ASXL1-MT cooperates with BAP1 to deubiquitinate and activate AKT. Overactive Akt/mTOR signaling induced by ASXL1-MT results in aberrant proliferation and dysfunction of HSCs associated with age-related accumulation of DNA damage. Treatment with an mTOR inhibitor rapamycin ameliorates aberrant expansion of the HSC compartment as well as dysregulated hematopoiesis in aged ASXL1-MT KI mice. Our findings suggest that ASXL1-MT provokes dysfunction of HSCs, whereas it confers clonal advantage on HSCs over time, leading to the development of CH. ASXL1 mutations are frequently found in age-related clonal haemaotopoiesis (CH), but how they drive CH is unclear. Here the authors show that expression of C-terminal truncated ASXL1 in haematopoietic stem cells (HSCs) leads to Akt de-ubiquitination, activated Akt/mTOR signaling, and aberrant HSC proliferation.

The most frequently mutated oncogene in cancer is KRAS , which uses alternative fourth exons to generate two gene products (KRAS4A and KRAS4B) that differ only in their C-terminal membrane-targeting region 1 . Because oncogenic mutations occur in exons 2 or 3, two constitutively active KRAS proteins—each capable of transforming cells—are encoded when KRAS is activated by mutation 2 . No functional distinctions among the splice variants have so far been established. Oncogenic KRAS alters the metabolism of tumour cells 3 in several ways, including increased glucose uptake and glycolysis even in the presence of abundant oxygen 4 (the Warburg effect). Whereas these metabolic effects of oncogenic KRAS have been explained by transcriptional upregulation of glucose transporters and glycolytic enzymes 3 – 5 , it is not known whether there is direct regulation of metabolic enzymes. Here we report a direct, GTP-dependent interaction between KRAS4A and hexokinase 1 (HK1) that alters the activity of the kinase, and thereby establish that HK1 is an effector of KRAS4A. This interaction is unique to KRAS4A because the palmitoylation–depalmitoylation cycle of this RAS isoform enables colocalization with HK1 on the outer mitochondrial membrane. The expression of KRAS4A in cancer may drive unique metabolic vulnerabilities that can be exploited therapeutically. KRAS4A interacts directly with hexokinase 1 in a GTP-dependent manner at the outer mitochondrial membrane, leading to kinase activation and an increase in glucose uptake and glycolysis in tumour cells.

Gasdermin E (GSDME/DFNA5) cleavage by caspase-3 liberates the GSDME-N domain, which mediates pyroptosis by forming pores in the plasma membrane. Here we show that GSDME-N also permeabilizes the mitochondrial membrane, releasing cytochrome c and activating the apoptosome. Cytochrome c release and caspase-3 activation in response to intrinsic and extrinsic apoptotic stimuli are significantly reduced in GSDME-deficient cells comparing with wild type cells. GSDME deficiency also accelerates cell growth in culture and in a mouse model of melanoma. Phosphomimetic mutation of the highly conserved phosphorylatable Thr6 residue of GSDME, inhibits its pore-forming activity, thus uncovering a potential mechanism by which GSDME might be regulated. Like GSDME-N, inflammasome-generated gasdermin D-N (GSDMD-N), can also permeabilize the mitochondria linking inflammasome activation to downstream activation of the apoptosome. Collectively, our results point to a role of gasdermin proteins in targeting the mitochondria to promote cytochrome c release to augment the mitochondrial apoptotic pathway. Gasdermins mediate lytic cell death by forming pores in the plasma membrane. Here the authors show that gasdermins also permeabilize mitochondrial membrane, thereby facilitating intrinsic apoptosis pathway, downstream of apoptotic (Gasdermin E) and inflammatory (Gasdermin D) caspase activation.

The role of the mitochondrial permeability transition pore (MPTP) in apoptosis of nucleated cells is well documented. In contrast, the role of MPTP in apoptosis of anucleated platelets is largely unknown. The aim of this study was to elucidate the contribution of MPTP in the control of different manifestations of platelet apoptosis by analyzing the effect of cyclosporin A (CsA), a potent inhibitor of MPTP formation. Using flow cytometry, we studied the effect of pretreatment of platelets with CsA on apoptotic responses in human platelets stimulated with calcium ionophore A23187. We found that CsA inhibited A23187-stimulated platelet apoptosis, completely preventing (i) depolarization of mitochondrial inner membrane potential (ΔΨm), (ii) activation of cytosolic apoptosis executioner caspase-3, (iii) platelet shrinkage, and (iv) fragmentation of platelets to microparticles, but (v) only partially (≈25%), inhibiting phosphatidylserine (PS) exposure on the platelet surface. This study shows that MPTP formation is upstream of ΔΨm depolarization, caspase-3 activation, platelet shrinkage and microparticle formation, and stringently controls these apoptotic events in A23187-stimulated platelets but is less involved in PS externalization. These data also indicate that CsA may rescue platelets from apoptosis, preventing caspase-3 activation and inhibiting the terminal cellular manifestations of platelet apoptosis, such as platelet shrinkage and degradation to microparticles. Furthermore, the results suggest a novel potentially useful application of CsA as an inhibitor of platelet demise through apoptosis in thrombocytopenias associated with enhanced platelet apoptosis.

The mitochondrial unfolded protein response (UPRmt) is characterized by the transcriptional induction of mitochondrial chaperone and protease genes in response to impaired mitochondrial proteostasis and is regulated by ATF5 and CHOP in mammalian cells. However, the detailed mechanisms underlying the UPRmt are currently unclear. Here, we show that HSF1 is required for activation of mitochondrial chaperone genes, including HSP60, HSP10, and mtHSP70, in mouse embryonic fibroblasts during inhibition of matrix chaperone TRAP1, protease Lon, or electron transfer complex 1 activity. HSF1 bound constitutively to mitochondrial chaperone gene promoters, and we observed that its occupancy was remarkably enhanced at different levels during the UPRmt. Furthermore, HSF1 supported the maintenance of mitochondrial function under the same conditions. These results demonstrate that HSF1 is required for induction of mitochondrial chaperones during the UPRmt, and thus, it may be one of the guardians of mitochondrial function under conditions of impaired mitochondrial proteostasis. The mitochondrial unfolded protein response (UPRmt) is characterized by the transcriptional induction of mitochondrial chaperone and protease genes in response to impaired mitochondrial proteostasis. Here, we show that heat shock transcription factor (HSF1) is required for activation of mitochondrial chaperone genes and supports the maintenance of mitochondrial function in mouse cells during the UPRmt.

Intrinsic apoptotic stimuli initiate mammalian cells’ apoptotic program by first activating the proteins that have only Bcl-2 homology domain 3 (BH3), such as Bcl-2 interacting mediator of cell death (Bim) and truncated BH3 interacting death domain agonist (tBid), which in turn trigger conformational changes in BCL2-associated X (Bax) and BCL2-antagonist/killer (Bak) proteins that enable oligomer formation on the mitochondria, causing cytochrome c and other apoptogenic proteins in the intermembrane space to leak out. Leaked cytochrome c then initiates apoptotic caspase activation through a well-defined biochemical pathway. However, how oligomerized Bax and Bak cause cytochrome c release from mitochondria remains unknown. We report here the establishment of cell lines in which Bim or tBid can be inducibly expressed to initiate apoptosis in a controlled, quantitative manner. We used these cell lines to examine apoptotic events after Bax and Bak oligomerization but before cytochrome c release. The mitochondrial metalloprotease OMA1 was activated in this system in a Bax- and Bak-dependent fashion. Activated OMA1 cleaved the dynamin-like GTPase, optical nerve atrophy 1, an event that is critical for remodeling of mitochondrial cristae. Knockdown or knockout of OMA1 in these cells attenuated cytochrome c release. Thus it is clear that oligomerized Bax and Bak trigger apoptosis by causing both the permeabilization of the mitochondrial outer membrane and activation OMA1. Significance The release of cytochrome c from its normal intermembrane space in mitochondria marks the initiation of apoptosis in mammalian cells. The process is triggered by the aggregation of B-cell leukemia/lymphoma 2 (BCL2)-associated X (Bax) and BCL2-antagonist/killer (Bak) proteins on the surface of mitochondria. We found that a mitochondrial inner membrane protease, OMA1 (overlapping activity with m -AAA protease), is specifically activated and is responsible for cleaving another inner membrane protein, optical nerve atrophy 1 (OPA1), upon Bax/Bak aggregation. The cleavage of OPA1 triggers the remodeling of mitochondrial cristae, allowing the majority of cytochrome c inside the cristae to be released. This finding provided a more comprehensive understanding of this critical molecular event during apoptosis.