Neuroferritinopathy is a rare genetic disease with a dominant autosomal transmission caused by mutations of the ferritin light chain gene (FTL). It belongs to Neurodegeneration with Brain Iron Accumulation, a group of disorders where iron dysregulation is tightly associated with neurodegeneration. We studied the 498-499InsTC mutation which causes the substitution of the last 9 amino acids and an elongation of extra 16 amino acids at the C-terminus of L-ferritin peptide. An analysis with cyclic voltammetry on the purified protein showed that this structural modification severely reduces the ability of the protein to store iron. In order to analyze the impact of the mutation in vivo, we generated mouse models for the some pathogenic human FTL gene in FVB and C57BL/6J strains. Transgenic mice in the FVB background showed high accumulation of the mutated ferritin in brain where it correlated with increased iron deposition with age, as scored by magnetic resonance imaging. Notably, the accumulation of iron-ferritin bodies was accompanied by signs of oxidative damage. In the C57BL/6 background, both the expression of the mutant ferritin and the iron levels were lower than in the FVB strain. Nevertheless, also these mice showed oxidative alterations in the brain. Furthermore, post-natal hippocampal neurons obtained from these mice experienced a marked increased cell death in response to chronic iron overload and/or acute oxidative stress, in comparison to wild-type neurons. Ultrastructural analyses revealed an accumulation of lipofuscin granules associated with iron deposits, particularly enriched in the cerebellum and striatum of our transgenic mice. Finally, experimental subjects were tested throughout development and aging at 2-, 8- and 18-months for behavioral phenotype. Rotarod test revealed a progressive impaired motor coordination building up with age, FTL mutant old mice showing a shorter latency to fall from the apparatus, according to higher accumulation of iron aggregates in the striatum. Our data show that our 498-499InsTC mouse models recapitulate early pathological and clinical traits of the human neuroferritinopathy, thus providing a valuable model for the study of the disease. Finally, we propose a mechanistic model of lipofuscine formation that can account for the etiopathogenesis of human neuroferritinopathy.

Mutations in the inositol 5-phosphatase OCRL cause Lowe syndrome and Dent's disease. Although OCRL, a direct clathrin interactor, is recruited to late-stage clathrin-coated pits, clinical manifestations have been primarily attributed to intracellular sorting defects. Here we show that OCRL loss in Lowe syndrome patient fibroblasts impacts clathrin-mediated endocytosis and results in an endocytic defect. These cells exhibit an accumulation of clathrin-coated vesicles and an increase in U-shaped clathrin-coated pits, which may result from sequestration of coat components on uncoated vesicles. Endocytic vesicles that fail to lose their coat nucleate the majority of the numerous actin comets present in patient cells. SNX9, an adaptor that couples late-stage endocytic coated pits to actin polymerization and which we found to bind OCRL directly, remains associated with such vesicles. These results indicate that OCRL acts as an uncoating factor and that defects in clathrin-mediated endocytosis likely contribute to pathology in patients with OCRL mutations.

FAM46C is a non-canonical poly(A) polymerase uniquely mutated in up to 20% of multiple myeloma (MM) patients, implying a tissue-specific tumor suppressor function. Here, we report that FAM46C selectively stabilizes mRNAs encoding endoplasmic reticulum (ER)-targeted proteins, thereby concertedly enhancing the expression of proteins that control ER protein import, folding, N-glycosylation, and trafficking and boosting protein secretion. This role requires the interaction with the ER membrane resident proteins FNDC3A and FNDC3B. In MM cells, FAM46C expression raises secretory capacity beyond sustainability, inducing ROS accumulation, ATP shortage, and cell death. FAM46C activity is regulated through rapid proteasomal degradation or the inhibitory interaction with the ZZ domain of the autophagic receptor p62 that hinders its association with FNDC3 proteins via sequestration in p62+ aggregates. Altogether, our data disclose a p62/FAM46C/FNDC3 circuit coordinating sustainable secretory activity and survival, providing an explanation for the MM-specific oncosuppressive role of FAM46C and uncovering potential therapeutic opportunities against cancer.

Dementia is a neurocognitive disorder characterized by a progressive memory loss and impairment in cognitive and functional abilities. Autophagy and mitophagy are two important cellular processes by which the damaged intracellular components are degraded by lysosomes. To investigate the contribution of autophagy and mitophagy in degenerative diseases, we investigated the serum levels of specific autophagic markers (ATG5 protein) and mitophagic markers (Parkin protein) in a population of older patients by enzyme-linked immunosorbent assay. Two hundred elderly (≥65 years) outpatients were included in the study: 40 (20 F and 20 M) with mild-moderate late onset Alzheimer's disease (AD); 40 (20 F and 20 M) affected by vascular dementia (VAD); 40 with mild cognitive impairment (MCI); 40 (20 F and 20 M) with "mixed" dementia (MD); 40 subjects without signs of cognitive impairment were included as sex-matched controls. Our data indicated that, in serum samples, ATG5 and Parkin were both elevated in controls, and that VAD compared with AD, MCI and MD (all p < 0.01). Patients affected by AD, MD, and MCI showed significantly reduced circulating levels of both ATG5 and Parkin compared to healthy controls and VAD individuals, reflecting a significant down-regulation of autophagy and mitophagy pathways in these groups of patients. The measurement of serum levels of ATG5 and Parkin may represent an easily accessible diagnostic tool for the early monitoring of patients with cognitive decline.

Tissue engineering-based bone graft is an emerging viable treatment modality to repair and regenerate tissues damaged as a result of diseases or injuries. The structure and composition of scaffolds should modulate the classical osteogenic pathways in human stem cells. The osteoinductivity properties of the hydroxylapatite-collagen hybrid scaffold named Coll/Pro Osteon 200 were investigated in an in vitro model of human adipose mesenchymal stem cells (hASCs), whereas the clinical evaluation was carried out in maxillofacial patients. Differentially expressed genes (DEGs) induced by the scaffold were analyzed using the Osteogenesis RT2 PCR Array. The osteoinductivity potential of the scaffold was also investigated by studying the alkaline phosphatase (ALP) activity, matrix mineralization, osteocalcin (OCN), and CLEC3B expression protein. Fifty patients who underwent zygomatic augmentation and bimaxillary osteotomy were evaluated clinically, radiologically, and histologically during a 3-year follow-up. Among DEGs, osteogenesis-related genes, including BMP1/2, ALP, BGLAP, SP7, RUNX2, SPP1, and EGFR, which play important roles in osteogenesis, were found to be upregulated. The genes to cartilage condensation SOX9, BMPR1B, and osteoclast cells TNFSF11 were detected upregulated at every time point of the investigation. This scaffold has a high osteoinductivity revealed by the matrix mineralization, ALP activity, OCN, and CLEC3B expression proteins. Clinical evaluation evidences that the biomaterial promotes bone regrowth. Histological results of biopsy specimens from patients showed prominent ossification. Experimental data using the Coll/Pro Osteon 200 indicate that clinical evaluation of bone regrowth in patients, after scaffold implantation, was supported by DEGs implicated in skeletal development as shown in "in vitro" experiments with hASCs.

The unfolded protein response (UPR) is one of the major cell-autonomous proteostatic stress responses. The UPR has been implicated in the pathogenesis of neurodegenerative diseases and is therefore actively investigated as therapeutic target. In this respect, cell non-autonomous effects of the UPR including the reported cell-to-cell transmission of UPR activity may be highly important. A pharmaca-based UPR induction was employed to generate conditioned media (CM) from CM-donating neuronal ('donor') cells (SK-N-SH and primary mouse neurons). As previously reported, upon subsequent transfer of CM to naive neuronal 'acceptor' cells, we confirmed UPR target mRNA and protein expression by qPCR and automated microscopy. However, UPR target gene expression was also induced in the absence of donor cells, indicating carry-over of pharmaca. Genetic induction of single pathways of the UPR in donor cells did not result in UPR transmission to acceptor cells. Moreover, no transmission was detected upon full UPR activation by nutrient deprivation or inducible expression of the heavy chain of immunoglobulin M in donor HeLa cells. In addition, in direct co-culture of donor cells expressing the immunoglobulin M heavy chain and fluorescent UPR reporter acceptor HeLa cells, UPR transmission was not observed. In conclusion, carry-over of pharmaca is a major confounding factor in pharmaca-based UPR transmission protocols that are therefore unsuitable to study cell-to-cell UPR transmission. In addition, the absence of UPR transmission in non-pharmaca-based models of UPR activation indicates that cell-to-cell UPR transmission does not occur in cell culture.

Adaptor protein 2 (AP2) is a major constituent of clathrin-coated pits (CCPs). Whether it is essential for all forms of clathrin-mediated endocytosis (CME) in mammalian cells is an open issue. Here, we demonstrate, by live TIRF microscopy, the existence of a subclass of relatively short-lived CCPs lacking AP2 under physiological, unperturbed conditions. This subclass is retained in AP2-knockout cells and is able to support the internalization of epidermal growth factor receptor (EGFR) but not of transferrin receptor (TfR). The AP2-independent internalization mechanism relies on the endocytic adaptors eps15, eps15L1, and epsin1. The absence of AP2 impairs the recycling of the EGFR to the cell surface, thereby augmenting its degradation. Accordingly, under conditions of AP2 ablation, we detected dampening of EGFR-dependent AKT signaling and cell migration, arguing that distinct classes of CCPs could provide specialized functions in regulating EGFR recycling and signaling.

Ferritin is the main intracellular protein of iron storage with a central role in the regulation of iron metabolism and detoxification. Nucleotide insertions in the last exon of the ferritin light chain cause a neurodegenerative disease known as Neuroferritinopathy, characterized by iron deposition in the brain, particularly in the cerebellum, basal ganglia and motor cortex. The disease progresses relentlessly, leading to dystonia, chorea, motor disability and neuropsychiatry features. The characterization of a good animal model is required to compare and contrast specific features with the human disease, in order to gain new insights on the consequences of chronic iron overload on brain function and behavior. To this aim we studied an animal model expressing the pathogenic human FTL mutant 498InsTC under the phosphoglycerate kinase (PGK) promoter. Transgenic (Tg) mice showed strong accumulation of the mutated protein in the brain, which increased with age, and this was accompanied by brain accumulation of ferritin/iron bodies, the main pathologic hallmark of human neuroferritinopathy. Tg-mice were tested throughout development and aging at 2-, 8- and 18-months for motor coordination and balance (Beam Walking and Footprint tests). The Tg-mice showed a significant decrease in motor coordination at 8 and 18 months of age, with a shorter latency to fall and abnormal gait. Furthermore, one group of aged naïve subjects was challenged with two herbicides (Paraquat and Maneb) known to cause oxidative damage. The treatment led to a paradoxical increase in behavioral activation in the transgenic mice, suggestive of altered functioning of the dopaminergic system. Overall, data indicate that mice carrying the pathogenic FTL498InsTC mutation show motor deficits with a developmental profile suggestive of a progressive pathology, as in the human disease. These mice could be a powerful tool to study the neurodegenerative mechanisms leading to the disease and help developing specific therapeutic targets.

It has been recently demonstrated that high pre-treatment levels of miR-29b positively correlated with the response of patients with acute myeloid leukaemia (AML) to hypomethylating agents. Upmodulation of miR-29b by restoring its transcriptional machinery appears indeed a tool to improve therapeutic response in AML. In cells from acute promyelocytic leukaemia (APL), miR-29b is regulated by PU.1, in turn upmodulated by agonists currently used to treat APL. We explored here the ability of PU.1 to also regulate miR-29b in non-APL cells, in order to identify agonists that, upmodulating PU.1 may be beneficial in hypomethylating agents-based therapies. We found that PU.1 may regulate miR-29b in the non-APL Kasumi-1 cells, showing the t(8;21) chromosomal rearrangement, which is prevalent in AML and correlated with a relatively low survival. We demonstrated that the PU.1-mediated contribution of the 2 miR-29b precursors is cell-related and almost completely dependent on adequate levels of Vav1. Nuclear PU.1/Vav1 association accompanies the transcription of miR-29b but, at variance with the APL-derived NB4 cells, in which the protein is required for the association of PU.1 with both miRNA promoters, Vav1 is part of molecular complexes to the PU.1 consensus site in Kasumi-1. Our results add new information on the transcriptional machinery that regulates miR-29b expression in AML-derived cells and may help in identifying drugs useful in upmodulation of this miRNA in pre-treatment of patients with non-APL leukaemia who can take advantage from hypomethylating agent-based therapies.

Depletion of liprin-α1, ERC1 or LL5 scaffolds inhibits extracellular matrix degradation by invasive cells. These proteins co-accumulate near invadosomes in NIH-Src cells, identifying a novel invadosome-associated compartment distinct from the core and adhesion ring of invadosomes. Depletion of either protein perturbs the organization of invadosomes without influencing the recruitment of MT1-MMP metalloprotease. Liprin-α1 is not required for de novo formation of invadosomes after their disassembly by microtubules and Src inhibitors, while its depletion inhibits invadosome motility, thus affecting matrix degradation. Fluorescence recovery after photobleaching shows that the invadosome-associated compartment is dynamic, while correlative light immunoelectron microscopy identifies bona fide membrane-free invadosome-associated regions enriched in liprin-α1, which is virtually excluded from the invadosome core. The results indicate that liprin-α1, LL5 and ERC1 define a novel dynamic membrane-less compartment that regulates matrix degradation by affecting invadosome motility.

The unfolded protein response (UPR) is an intracellular signaling pathway that counteracts variable stresses that impair protein folding in the endoplasmic reticulum (ER). As such, the UPR is thought to be a homeostat that finely tunes ER protein folding capacity and ER abundance according to need. The mechanism by which the ER stress sensor Ire1 is activated by unfolded proteins and the role that the ER chaperone protein BiP plays in Ire1 regulation have remained unclear. Here we show that the UPR matches its output to the magnitude of the stress by regulating the duration of Ire1 signaling. BiP binding to Ire1 serves to desensitize Ire1 to low levels of stress and promotes its deactivation when favorable folding conditions are restored to the ER. We propose that, mechanistically, BiP achieves these functions by sequestering inactive Ire1 molecules, thereby providing a barrier to oligomerization and activation, and a stabilizing interaction that facilitates de-oligomerization and deactivation. Thus BiP binding to or release from Ire1 is not instrumental for switching the UPR on and off as previously posed. By contrast, BiP provides a buffer for inactive Ire1 molecules that ensures an appropriate response to restore protein folding homeostasis to the ER by modulating the sensitivity and dynamics of Ire1 activity.

Clathrin light chains (CLCa and CLCb) are major constituents of clathrin-coated vesicles. Unique functions for these evolutionary conserved paralogs remain elusive, and their role in clathrin-mediated endocytosis in mammalian cells is debated. Here, we find and structurally characterize a direct and selective interaction between CLCa and the long isoform of the actin motor protein myosin VI, which is expressed exclusively in highly polarized tissues. Using genetically-reconstituted Caco-2 cysts as proxy for polarized epithelia, we provide evidence for coordinated action of myosin VI and CLCa at the apical surface where these proteins are essential for fission of clathrin-coated pits. We further find that myosin VI and Huntingtin-interacting protein 1-related protein (Hip1R) are mutually exclusive interactors with CLCa, and suggest a model for the sequential function of myosin VI and Hip1R in actin-mediated clathrin-coated vesicle budding.

Disorders of lysosomal physiology have increasingly been found to underlie the pathology of a rapidly growing cast of neurodevelopmental disorders and sporadic diseases of aging. One cardinal aspect of lysosomal (dys)function is lysosomal acidification in which defects trigger lysosomal stress signaling and defects in proteolytic capacity. We have developed a genetically encoded ratiometric probe to measure lysosomal pH coupled with a purification tag to efficiently purify lysosomes for both proteomic and in vitro evaluation of their function. Using our probe, we showed that lysosomal pH is remarkably stable over a period of days in a variety of cell types. Additionally, this probe can be used to determine that lysosomal stress signaling via TFEB is uncoupled from gross changes in lysosomal pH. Finally, we demonstrated that while overexpression of ARL8B GTPase causes striking alkalinization of peripheral lysosomes in HEK293 T cells, peripheral lysosomes per se are no less acidic than juxtanuclear lysosomes in our cell lines.Abbreviations: ARL8B: ADP ribosylation factor like GTPase 8B; ATP: adenosine triphosphate; ATP5F1B/ATPB: ATP synthase F1 subunit beta; ATP6V1A: ATPase H+ transporting V1 subunit A; Baf: bafilomycin A1; BLOC-1: biogenesis of lysosome-related organelles complex 1; BSA: bovine serum albumin; Cos7: African green monkey kidney fibroblast-like cell line; CQ: chloroquine; CTSB: cathepsin B; CYCS: cytochrome c, somatic; DAPI: 4',6-diamidino -2- phenylindole; DIC: differential interference contrast; DIV: days in vitro; DMEM: Dulbecco's modified Eagle's medium;‎ E8: embryonic day 8; EEA1: early endosome antigen 1; EGTA: ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid; ER: endoplasmic reticulum; FBS: fetal bovine serum; FITC: fluorescein isothiocyanate; GABARAPL2: GABA type A receptor associated protein like 2; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GOLGA2/GM130: golgin A2; GTP: guanosine triphosphate; HEK293T: human embryonic kidney 293 cells, that expresses a mutant version of the SV40 large T antigen; HeLa: Henrietta Lacks-derived cell; HEPES: 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; HRP: horseradish peroxidase; IGF2R/ciM6PR: insulin like growth factor 2 receptor; LAMP1/2: lysosomal associated membrane protein 1/2; LMAN2/VIP36: lectin, mannose binding 2; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MTORC1: mechanistic target of rapamycin kinase complex 1; PCR: polymerase chain reaction; PDL: poly-d-lysine; PGK1p: promotor from human phosphoglycerate kinase 1; PIKFYVE: phosphoinositide kinase, FYVE-type zinc finger containing; PPT1/CLN1: palmitoyl-protein thioesterase 1; RPS6KB1/p70: ribosomal protein S6 kinase B1; STAT3: signal transducer and activator of transcription 3; TAX1BP1: Tax1 binding protein 1; TFEB: transcription factor EB; TGN: trans-Golgi network; TGOLN2/TGN46: trans-Golgi network protein 2; TIRF: total internal reflection fluorescence; TMEM106B: transmembrane protein 106B; TOR: target of rapamycin; TRPM2: transient receptor potential cation channel subfamily M member 2; V-ATPase: vacuolar-type proton-translocating ATPase; VPS35: VPS35 retromer complex component.

Abnormal levels of pyruvate and lactate were reported in multiple sclerosis (MS). We studied the response of markers of mitochondrial function to rehabilitation in relation to type, intensity and endurance performance in severely disabled MS patients. Forty-six progressive MS patients were randomized to receive 12 walking sessions of robot-assisted gait training (RAGT, n = 23) or conventional overground therapy (CT, n = 23). Ten healthy subjects were also studied. Blood samples were collected to determine lactate, pyruvate, and glutathione levels and lactate/pyruvate ratio pre-post rehabilitation. In vivo muscle metabolism and endurance walking capacity were assessed by resting muscle oxygen consumption (rmVO2) using near-infrared spectroscopy and by six-minute walking distance (6MWD), respectively. The levels of mitochondrial biomarkers and rmVO2, altered at baseline with respect to healthy subjects, improved after rehabilitation in the whole population. In the two groups, an enhanced response was observed after RAGT compared to CT for lactate (p = 0.012), glutathione (<0.001), lactate/pyruvate ratio (p = 0.08) and rmVO2 (p = 0.07). Metabolic biomarkers and 6MWD improvements were exclusively correlated with a training speed markedly below individual gait speed. In severely disabled MS patients, rehabilitation rebalanced altered serum metabolic and muscle parameters, with RAGT being more effective than CT. A determinable slow training speed was associated with better metabolic and functional recovery. Trial Registration: ClinicalTrials.gov NCT02421731.

Leber's hereditary optic neuropathy (LHON), a disease associated with a mitochondrial DNA mutation, is characterized by blindness due to degeneration of retinal ganglion cells (RGCs) and their axons, which form the optic nerve. We show that a sustained pathological autophagy and compartment-specific mitophagy activity affects LHON patient-derived cells and cybrids, as well as induced pluripotent-stem-cell-derived neurons. This is variably counterbalanced by compensatory mitobiogenesis. The aberrant quality control disrupts mitochondrial homeostasis as reflected by defective bioenergetics and excessive reactive oxygen species production, a stress phenotype that ultimately challenges cell viability by increasing the rate of apoptosis. We counteract this pathological mechanism by using autophagy regulators (clozapine and chloroquine) and redox modulators (idebenone), as well as genetically activating mitochondrial biogenesis (PGC1-α overexpression). This study substantially advances our understanding of LHON pathophysiology, providing an integrated paradigm for pathogenesis of mitochondrial diseases and druggable targets for therapy.

The precise molecular mechanisms that coordinate apoptosis and autophagy in cancer remain to be determined. Here, we provide evidence that the tumor suppressor promyelocytic leukemia protein (PML) controls autophagosome formation at mitochondria-associated membranes (MAMs) and, thus, autophagy induction. Our in vitro and in vivo results demonstrate how PML functions as a repressor of autophagy. PML loss promotes tumor development, providing a growth advantage to tumor cells that use autophagy as a cell survival strategy during stress conditions. These findings demonstrate that autophagy inhibition could be paired with a chemotherapeutic agent to develop anticancer strategies for tumors that present PML downregulation.

Triplication of the SNCA gene, encoding the protein alpha-Synuclein (αSyn), is a rare cause of aggressive and early-onset parkinsonism. Herein, we generated iPSCs from two siblings with a recently described compact SNCA gene triplication and suffering from severe motor impairments, psychiatric symptoms, and cognitive deterioration. Using CRISPR/Cas9 gene editing, each SNCA copy was inactivated by targeted indel mutations generating a panel of isogenic iPSCs with a decremental number from 4 down to none of functional SNCA gene alleles. We differentiated these iPSC lines in midbrain dopaminergic (DA) neuronal cultures to characterize αSyn aggregation in native and seeded conditions and evaluate its associated cellular dysfunctions. Utilizing a new nanobody-based biosensor combined with super-resolved imaging, we were able to visualize and measure αSyn aggregates in early DA neurons in unstimulated conditions. Calcium dysregulation and mitochondrial alterations were the first pathological signs detectable in early differentiated DA neuronal cultures. Accelerated αSyn aggregation was induced by exposing neurons to structurally well-characterized synthetic αSyn fibrils. 4xSNCA DA neurons showed the highest vulnerability, which was associated with high levels of oxidized DA and amplified by TAX1BP1 gene disruption. Seeded DA neurons developed large αSyn deposits whose morphology and internal constituents resembled Lewy bodies commonly observed in Parkinson's disease (PD) patient brain tissues. These findings provide strong evidence that this isogenic panel of iPSCs with SNCA multiplications offers a remarkable cellular platform to investigate mechanisms of PD and validate candidate inhibitors of native and seeded αSyn aggregation.

In mouse liver hepatocytes, nearly half of the surface area of every mitochondrion is covered by wrappER, a wrapping-type of ER that is rich in fatty acids and synthesizes lipoproteins (VLDL) (Anastasia et al. in Cell Rep 34:108873, 2021; Hurtley in Science (80- ) 372:142-143, 2021; Ilacqua et al. in J Cell Sci 135:1-11, 2021). A disruption of the ultrastructure of the wrappER-mitochondria contact results in altered fatty acid flux, leading to hepatic dyslipidemia (Anastasia et al. 2021). The molecular mechanism that regulates the extent of wrappER-mitochondria contacts is unknown.

EPS15 and its homologous EPS15L1 are endocytic accessory proteins. Studies in mammalian cell lines suggested that EPS15 and EPS15L1 regulate endocytosis in a redundant manner. However, at the organismal level, it is not known to which extent the functions of the two proteins overlap. Here, by exploiting various constitutive and conditional null mice, we report redundant and nonredundant functions of the two proteins. EPS15L1 displays a unique nonredundant role in the nervous system, whereas both proteins are fundamental during embryo development as shown by the embryonic lethality of -Eps15/Eps15L1-double KO mice. At the cellular level, the major process redundantly regulated by EPS15 and EPS15L1 is the endocytosis of the transferrin receptor, a pathway that sustains the development of red blood cells and controls iron homeostasis. Consequently, hematopoietic-specific conditional Eps15/Eps15L1-double KO mice display traits of microcytic hypochromic anemia, due to a cell-autonomous defect in iron internalization.

Mounting evidence links neurodegenerative disorders such as Parkinson disease and Alzheimer disease with mitochondrial dysfunction, and recent emphasis has focused on mitochondrial dynamics and quality control. Mitochondrial dynamics and mtDNA maintenance is another link recently emerged, implicating mutations in the mitochondrial fusion genes OPA1 and MFN2 in the pathogenesis of multisystem syndromes characterized by neurodegeneration and accumulation of mtDNA multiple deletions in postmitotic tissues. Here, we report 2 Italian families affected by dominant chronic progressive external ophthalmoplegia (CPEO) complicated by parkinsonism and dementia.