Loss of Mfn1 but not Mfn2 enhances adipogenesis.

Objective: A biallelic missense mutation in mitofusin 2 (MFN2) causes multiple symmetric lipomatosis and partial lipodystrophy, implicating disruption of mitochondrial fusion or interaction with other organelles in adipocyte differentiation, growth and/or survival. In this study, we aimed to document the impact of loss of mitofusin 1 (Mfn1) or 2 (Mfn2) on adipogenesis in cultured cells.

Methods: We characterised adipocyte differentiation of wildtype (WT), Mfn1-/- and Mfn2-/- mouse embryonic fibroblasts (MEFs) and 3T3-L1 preadipocytes in which Mfn1 or 2 levels were reduced using siRNA.

Molecular mechanisms of mitochondrial dynamics.

Mitochondria not only synthesize energy required for cellular functions but are also involved in numerous cellular pathways including apoptosis, calcium homoeostasis, inflammation and immunity. Mitochondria are dynamic organelles that undergo cycles of fission and fusion, and these transitions between fragmented and hyperfused networks ensure mitochondrial function, enabling adaptations to metabolic changes or cellular stress. Defects in mitochondrial morphology have been associated with numerous diseases, highlighting the importance of elucidating the molecular mechanisms regulating mitochondrial morphology.

Distinct roles for the domains of the mitochondrial aspartate/glutamate carrier citrin in organellar localization and substrate transport.

Objective: Citrin, the mitochondrial aspartate/glutamate carrier isoform 2 (AGC2), is structurally and mechanistically the most complex SLC25 family member, because it consists of three domains and forms a homo-dimer. Each protomer has an N-terminal calcium-binding domain with EF-hands, followed by a substrate-transporting carrier domain and a C-terminal domain with an amphipathic helix. The absence or dysfunction of citrin leads to citrin deficiency, a highly prevalent pan-ethnic mitochondrial disease.

Biallelic PTPMT1 variants disrupt cardiolipin metabolism and lead to a neurodevelopmental syndrome.

Primary mitochondrial diseases (PMDs) are among the most common inherited neurological disorders. They are caused by pathogenic variants in mitochondrial or nuclear DNA that disrupt mitochondrial structure and/or function, leading to impaired oxidative phosphorylation (OXPHOS). One emerging subcategory of PMDs involves defective phospholipid (PL) metabolism.

MTFP1 controls mitochondrial fusion to regulate inner membrane quality control and maintain mtDNA levels.

Mitochondrial dynamics play a critical role in cell fate decisions and in controlling mtDNA levels and distribution. However, the molecular mechanisms linking mitochondrial membrane remodeling and quality control to mtDNA copy number (CN) regulation remain elusive. Here, we demonstrate that the inner mitochondrial membrane (IMM) protein mitochondrial fission process 1 (MTFP1) negatively regulates IMM fusion.

Preserved striatal innervation maintains motor function despite severe loss of nigral dopaminergic neurons.

Degeneration of dopaminergic neurons in the substantia nigra and their striatal axon terminals causes cardinal motor symptoms of Parkinson's disease. In idiopathic cases, high levels of mitochondrial DNA alterations, leading to mitochondrial dysfunction, are a central feature of these vulnerable neurons. Here we present a mouse model expressing the K320E variant of the mitochondrial helicase Twinkle in dopaminergic neurons, leading to accelerated mitochondrial DNA mutations.

Activity-dependent compartmentalization of dendritic mitochondria morphology through local regulation of fusion-fission balance in neurons in vivo.

Neuronal mitochondria play important roles beyond ATP generation, including Ca uptake, and therefore have instructive roles in synaptic function and neuronal response properties. Mitochondrial morphology differs significantly between the axon and dendrites of a given neuronal subtype, but in CA1 pyramidal neurons (PNs) of the hippocampus, mitochondria within the dendritic arbor also display a remarkable degree of subcellular, layer-specific compartmentalization. In the dendrites of these neurons, mitochondria morphology ranges from highly fused and elongated in the apical tuft, to more fragmented in the apical oblique and basal dendritic compartments, and thus occupy a smaller fraction of dendritic volume than in the apical tuft.

ESYT1 tethers the ER to mitochondria and is required for mitochondrial lipid and calcium homeostasis.

Mitochondria interact with the ER at structurally and functionally specialized membrane contact sites known as mitochondria-ER contact sites (MERCs). Combining proximity labelling (BioID), co-immunoprecipitation, confocal microscopy and subcellular fractionation, we found that the ER resident SMP-domain protein ESYT1 was enriched at MERCs, where it forms a complex with the outer mitochondrial membrane protein SYNJ2BP. BioID analyses using ER-targeted, outer mitochondrial membrane-targeted, and MERC-targeted baits, confirmed the presence of this complex at MERCs and the specificity of the interaction.

mtFociCounter for automated single-cell mitochondrial nucleoid quantification and reproducible foci analysis.

Mitochondrial DNA (mtDNA) encodes the core subunits for OXPHOS, essential in near-all eukaryotes. Packed into distinct foci (nucleoids) inside mitochondria, the number of mtDNA copies differs between cell-types and is affected in several human diseases. Currently, common protocols estimate per-cell mtDNA-molecule numbers by sequencing or qPCR from bulk samples.

Incorporating a Polyethyleneglycol Linker to Enhance the Hydrophilicity of Mitochondria-Targeted Triphenylphosphonium Constructs.

The targeting of bioactive molecules and probes to mitochondria can be achieved by coupling to the lipophilic triphenyl phosphonium (TPP) cation, which accumulates several hundred-fold within mitochondria in response to the mitochondrial membrane potential (Δψ ). Typically, a simple alkane links the TPP to its "cargo", increasing overall hydrophobicity. As it would be beneficial to enhance the water solubility of mitochondria-targeted compounds we explored the effects of replacing the alkyl linker with a polyethylene glycol (PEG).

Fumarate induces vesicular release of mtDNA to drive innate immunity.

Mutations in fumarate hydratase (FH) cause hereditary leiomyomatosis and renal cell carcinoma. Loss of FH in the kidney elicits several oncogenic signalling cascades through the accumulation of the oncometabolite fumarate. However, although the long-term consequences of FH loss have been described, the acute response has not so far been investigated.

A mouse model of human mitofusin-2-related lipodystrophy exhibits adipose-specific mitochondrial stress and reduced leptin secretion.

Mitochondrial dysfunction has been reported in obesity and insulin resistance, but primary genetic mitochondrial dysfunction is generally not associated with these, arguing against a straightforward causal relationship. A rare exception, recently identified in humans, is a syndrome of lower body adipose loss, leptin-deficient severe upper body adipose overgrowth, and insulin resistance caused by the p.Arg707Trp mutation in , encoding mitofusin 2.

Selective and reversible disruption of mitochondrial inner membrane protein complexes by lipophilic cations.

Triphenylphosphonium (TPP) derivatives are commonly used to target chemical into mitochondria. We show that alkyl-TPP cause reversible, dose- and hydrophobicity-dependent alterations of mitochondrial morphology and function and a selective decrease of mitochondrial inner membrane proteins including subunits of the respiratory chain complexes, as well as components of the mitochondrial calcium uniporter complex. The treatment with alkyl-TPP resulted in the cleavage of the pro-fusion and cristae organisation regulator Optic atrophy-1.

AMPK-dependent phosphorylation of MTFR1L regulates mitochondrial morphology.

Mitochondria are dynamic organelles that undergo membrane remodeling events in response to metabolic alterations to generate an adequate mitochondrial network. Here, we investigated the function of mitochondrial fission regulator 1-like protein (MTFR1L), an uncharacterized protein that has been identified in phosphoproteomic screens as a potential AMP-activated protein kinase (AMPK) substrate. We showed that MTFR1L is an outer mitochondrial membrane-localized protein modulating mitochondrial morphology.

Rapid and selective generation of HS within mitochondria protects against cardiac ischemia-reperfusion injury.

Mitochondria-targeted HS donors are thought to protect against acute ischemia-reperfusion (IR) injury by releasing HS that decreases oxidative damage. However, the rate of HS release by current donors is too slow to be effective upon administration following reperfusion. To overcome this limitation here we develop a mitochondria-targeted agent, MitoPerSulf that very rapidly releases HS within mitochondria.

Decreasing pdzd8-mediated mito-ER contacts improves organismal fitness and mitigates Aβ toxicity.

Mitochondria-ER contact sites (MERCs) orchestrate many important cellular functions including regulating mitochondrial quality control through mitophagy and mediating mitochondrial calcium uptake. Here, we identify and functionally characterize the Drosophila ortholog of the recently identified mammalian MERC protein, Pdzd8. We find that reducing pdzd8-mediated MERCs in neurons slows age-associated decline in locomotor activity and increases lifespan in Drosophila.

TMEM63C mutations cause mitochondrial morphology defects and underlie hereditary spastic paraplegia.

The hereditary spastic paraplegias (HSP) are among the most genetically diverse of all Mendelian disorders. They comprise a large group of neurodegenerative diseases that may be divided into 'pure HSP' in forms of the disease primarily entailing progressive lower-limb weakness and spasticity, and 'complex HSP' when these features are accompanied by other neurological (or non-neurological) clinical signs. Here, we identified biallelic variants in the transmembrane protein 63C (TMEM63C) gene, encoding a predicted osmosensitive calcium-permeable cation channel, in individuals with hereditary spastic paraplegias associated with mild intellectual disability in some, but not all cases.

Mitochondrial matrix-localized Src kinase regulates mitochondrial morphology.

The architecture of mitochondria adapts to physiological contexts: while mitochondrial fragmentation is usually associated to quality control and cell death, mitochondrial elongation often enhances cell survival during stress. Understanding how these events are regulated is important to elucidate how mitochondrial dynamics control cell fate. Here, we show that the tyrosine kinase Src regulates mitochondrial morphology.

Light-mediated discovery of surfaceome nanoscale organization and intercellular receptor interaction networks.

The molecular nanoscale organization of the surfaceome is a fundamental regulator of cellular signaling in health and disease. Technologies for mapping the spatial relationships of cell surface receptors and their extracellular signaling synapses would unlock theranostic opportunities to target protein communities and the possibility to engineer extracellular signaling. Here, we develop an optoproteomic technology termed LUX-MS that enables the targeted elucidation of acute protein interactions on and in between living cells using light-controlled singlet oxygen generators (SOG).

Quantifying inter-organelle membrane contact sites using proximity ligation assay in fixed optic nerve sections.

Membrane contact sites (MCS) play crucial roles in cell physiology with dysfunction in several MCS proteins being linked with neurological and optic nerve diseases. Although there have been significant advances in imaging these interactions over the past two decades with advanced electron microscopy techniques, super-resolution imaging and proximity-dependent fluorescent reporters, a technique to observe and quantify MCS in mammalian optic nerve tissues has not yet been reported. We demonstrate for the first time that proximity ligation assay (PLA), a technique already used in mammalian cell lines, can be used as an efficient method of quantifying inter-organelle contact sites, namely mitochondria-endoplasmic reticulum (ER) and mitochondria-late-endosomes, in mammalian optic nerve tissues treated with adeno-associated virus (AAV) gene therapy with wild-type or phosphomimetic (active) protrudin.

Mitochondrial translation is required for sustained killing by cytotoxic T cells.

T cell receptor activation of naïve CD8 T lymphocytes initiates their maturation into effector cytotoxic T lymphocytes (CTLs), which can kill cancer and virally infected cells. Although CTLs show an increased reliance on glycolysis upon acquisition of effector function, we found an essential requirement for mitochondria in target cell–killing. Acute mitochondrial depletion in USP30 (ubiquitin carboxyl-terminal hydrolase 30)–deficient CTLs markedly diminished killing capacity, although motility, signaling, and secretion were all intact.