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Frontiers in Molecular Neuroscience 2023Microglial cells ensure essential roles in brain homeostasis. In pathological condition, microglia adopt a common signature, called disease-associated microglial (DAM)...
Microglial cells ensure essential roles in brain homeostasis. In pathological condition, microglia adopt a common signature, called disease-associated microglial (DAM) signature, characterized by the loss of homeostatic genes and the induction of disease-associated genes. In X-linked adrenoleukodystrophy (X-ALD), the most common peroxisomal disease, microglial defect has been shown to precede myelin degradation and may actively contribute to the neurodegenerative process. We previously established BV-2 microglial cell models bearing mutations in peroxisomal genes that recapitulate some of the hallmarks of the peroxisomal β-oxidation defects such as very long-chain fatty acid (VLCFA) accumulation. In these cell lines, we used RNA-sequencing and identified large-scale reprogramming for genes involved in lipid metabolism, immune response, cell signaling, lysosome and autophagy, as well as a DAM-like signature. We highlighted cholesterol accumulation in plasma membranes and observed autophagy patterns in the cell mutants. We confirmed the upregulation or downregulation at the protein level for a few selected genes that mostly corroborated our observations and clearly demonstrated increased expression and secretion of DAM proteins in the BV-2 mutant cells. In conclusion, the peroxisomal defects in microglial cells not only impact on VLCFA metabolism but also force microglial cells to adopt a pathological phenotype likely representing a key contributor to the pathogenesis of peroxisomal disorders.
PubMed: 37138705
DOI: 10.3389/fnmol.2023.1170313 -
Brain : a Journal of Neurology Apr 2024The acyl-CoA-binding domain-containing protein 6 (ACBD6) is ubiquitously expressed, plays a role in the acylation of lipids and proteins and regulates the...
The acyl-CoA-binding domain-containing protein 6 (ACBD6) is ubiquitously expressed, plays a role in the acylation of lipids and proteins and regulates the N-myristoylation of proteins via N-myristoyltransferase enzymes (NMTs). However, its precise function in cells is still unclear, as is the consequence of ACBD6 defects on human pathophysiology. Using exome sequencing and extensive international data sharing efforts, we identified 45 affected individuals from 28 unrelated families (consanguinity 93%) with bi-allelic pathogenic, predominantly loss-of-function (18/20) variants in ACBD6. We generated zebrafish and Xenopus tropicalis acbd6 knockouts by CRISPR/Cas9 and characterized the role of ACBD6 on protein N-myristoylation with myristic acid alkyne (YnMyr) chemical proteomics in the model organisms and human cells, with the latter also being subjected further to ACBD6 peroxisomal localization studies. The affected individuals (23 males and 22 females), aged 1-50 years, typically present with a complex and progressive disease involving moderate-to-severe global developmental delay/intellectual disability (100%) with significant expressive language impairment (98%), movement disorders (97%), facial dysmorphism (95%) and mild cerebellar ataxia (85%) associated with gait impairment (94%), limb spasticity/hypertonia (76%), oculomotor (71%) and behavioural abnormalities (65%), overweight (59%), microcephaly (39%) and epilepsy (33%). The most conspicuous and common movement disorder was dystonia (94%), frequently leading to early-onset progressive postural deformities (97%), limb dystonia (55%) and cervical dystonia (31%). A jerky tremor in the upper limbs (63%), a mild head tremor (59%), parkinsonism/hypokinesia developing with advancing age (32%) and simple motor and vocal tics were among other frequent movement disorders. Midline brain malformations including corpus callosum abnormalities (70%), hypoplasia/agenesis of the anterior commissure (66%), short midbrain and small inferior cerebellar vermis (38% each) as well as hypertrophy of the clava (24%) were common neuroimaging findings. Acbd6-deficient zebrafish and Xenopus models effectively recapitulated many clinical phenotypes reported in patients including movement disorders, progressive neuromotor impairment, seizures, microcephaly, craniofacial dysmorphism and midbrain defects accompanied by developmental delay with increased mortality over time. Unlike ACBD5, ACBD6 did not show a peroxisomal localization and ACBD6-deficiency was not associated with altered peroxisomal parameters in patient fibroblasts. Significant differences in YnMyr-labelling were observed for 68 co- and 18 post-translationally N-myristoylated proteins in patient-derived fibroblasts. N-myristoylation was similarly affected in acbd6-deficient zebrafish and X. tropicalis models, including Fus, Marcks and Chchd-related proteins implicated in neurological diseases. The present study provides evidence that bi-allelic pathogenic variants in ACBD6 lead to a distinct neurodevelopmental syndrome accompanied by complex and progressive cognitive and movement disorders.
Topics: Animals; Female; Humans; Male; ATP-Binding Cassette Transporters; Intellectual Disability; Microcephaly; Movement Disorders; Nervous System Malformations; Neurodevelopmental Disorders; Tremor; Zebrafish; Infant; Child, Preschool; Child; Adolescent; Young Adult; Adult; Middle Aged
PubMed: 37951597
DOI: 10.1093/brain/awad380 -
Biomedicine & Pharmacotherapy =... Nov 2021X-linked adrenoleukodystrophy (X-ALD) is an inherited neurodegenerative disorder associated with mutations of the ABCD1 gene that encodes a peroxisomal transmembrane... (Review)
Review
X-linked adrenoleukodystrophy (X-ALD) is an inherited neurodegenerative disorder associated with mutations of the ABCD1 gene that encodes a peroxisomal transmembrane protein. It results in accumulation of very long chain fatty acids in tissues and body fluid. Along with other factors such as epigenetic and environmental involvement, ABCD1 mutation-provoked disorders can present different phenotypes including cerebral adrenoleukodystrophy (cALD), adrenomyeloneuropathy (AMN), and peripheral neuropathy. cALD is the most severe form that causes death in young childhood. Bone marrow transplantation and hematopoietic stem cell gene therapy are only effective when performed at an early stage of onsets in cALD. Nonetheless, current research and development of novel therapies are hampered by a lack of in-depth understanding disease pathophysiology and a lack of reliable cALD models. The Abcd1 and Abcd1/Abcd2 knock-out mouse models as well as the deficiency of Abcd1 rabbit models created in our lab, do not develop cALD phenotypes observed in human beings. In this review, we summarize the clinical and biochemical features of X-ALD, the progress of pre-clinical and clinical studies. Challenges and perspectives for future X-ALD studies are also discussed.
Topics: ATP Binding Cassette Transporter, Subfamily D; ATP Binding Cassette Transporter, Subfamily D, Member 1; Adrenoleukodystrophy; Animals; Disease Models, Animal; Genetic Predisposition to Disease; Humans; Mice, Knockout; Mutation; Phenotype; Prognosis; Rabbits; Species Specificity; Mice
PubMed: 34560537
DOI: 10.1016/j.biopha.2021.112214 -
Oxidative Medicine and Cellular... 2020Riboflavin transporter deficiency (RTD) is a childhood-onset neurodegenerative disorder characterized by progressive pontobulbar palsy, sensory and motor neuron...
Riboflavin transporter deficiency (RTD) is a childhood-onset neurodegenerative disorder characterized by progressive pontobulbar palsy, sensory and motor neuron degeneration, sensorineural hearing loss, and optic atrophy. As riboflavin (RF) is the precursor of FAD and FMN, we hypothesize that both mitochondrial and peroxisomal energy metabolism pathways involving flavoproteins could be directly affected in RTD, thus impacting cellular redox status. In the present work, we used induced pluripotent stem cells (iPSCs) from RTD patients to investigate morphofunctional features, focusing on mitochondrial and peroxisomal compartments. Using this model, we document the following RTD-associated alterations: (i) abnormal colony-forming ability and loss of cell-cell contacts, revealed by light, electron, and confocal microscopy, using tight junction marker ZO-1; (ii) mitochondrial ultrastructural abnormalities, involving shape, number, and intracellular distribution of the organelles, as assessed by focused ion beam/scanning electron microscopy (FIB/SEM); (iii) redox imbalance, with high levels of superoxide anion, as assessed by MitoSOX assay accompanied by abnormal mitochondrial polarization state, evaluated by JC-1 staining; (iv) altered immunofluorescence expression of antioxidant systems, namely, glutathione, superoxide dismutase 1 and 2, and catalase, as assessed by quantitatively evaluated confocal microscopy; and (v) peroxisomal downregulation, as demonstrated by levels and distribution of fatty acyl -oxidation enzymes. RF supplementation results in amelioration of cell phenotype and rescue of redox status, which was associated to improved ultrastructural features of mitochondria, thus strongly supporting patient treatment with RF, to restore mitochondrial- and peroxisomal-related aspects of energy dysmetabolism and oxidative stress in RTD syndrome.
Topics: Antioxidants; Base Sequence; Benzimidazoles; Biological Transport; Carbocyanines; Cell Shape; Child; Child, Preschool; Energy Metabolism; Humans; Induced Pluripotent Stem Cells; Mitochondria; Oxidation-Reduction; Peroxisomes; Receptors, G-Protein-Coupled; Riboflavin
PubMed: 32855765
DOI: 10.1155/2020/6821247 -
The FEBS Journal Nov 2019The leukodystrophies are a family of heritable disorders characterised by white matter degeneration, accompanied by variable clinical symptoms including loss of motor... (Review)
Review
The leukodystrophies are a family of heritable disorders characterised by white matter degeneration, accompanied by variable clinical symptoms including loss of motor function and cognitive decline. Now thought to include over 50 distinct disorders, there are a vast array of mechanisms underlying the pathology of these monogenic conditions and, accordingly, a range of animal models relating to each disorder. While both murine and zebrafish models continue to aid in the development of potential therapies, many of these models fail to truly recapitulate the human condition - thus leaving substantial weaknesses in our understanding of leukodystrophy pathogenesis. Additionally, the heterogeneity in leukodystrophy presentation - both in patients and in vivo models - often results in a narrow focus on single disorders in isolation across much of the literature. Thus, this review aims to synthesise prominent research regarding the most common leukodystrophies in order to provide an overview of key animal models and their utility in developing novel treatments. We begin by discussing the ongoing revolution across the leukodystrophy field following the rise of next generation sequencing, before focusing more extensively on existing animal models from the mouse and zebrafish fields. Finally, we explore how these preclinical models have shaped the development of therapeutic strategies currently in development. We propose future directions for the field and suggest a more critical view of the dogma which has underpinned leukodystrophy research for decades.
Topics: Adrenoleukodystrophy; Alexander Disease; Animals; Autoimmune Diseases of the Nervous System; Cognitive Dysfunction; Disease Models, Animal; Humans; Leukodystrophy, Globoid Cell; Leukodystrophy, Metachromatic; Leukoencephalopathies; Mice; Nervous System Malformations; Zebrafish
PubMed: 31520449
DOI: 10.1111/febs.15060 -
ELife Nov 2022Adrenoleukodystrophy protein (ALDP) is responsible for the transport of very-long-chain fatty acids (VLCFAs) and corresponding CoA-esters across the peroxisomal...
Adrenoleukodystrophy protein (ALDP) is responsible for the transport of very-long-chain fatty acids (VLCFAs) and corresponding CoA-esters across the peroxisomal membrane. Dysfunction of ALDP leads to peroxisomal metabolic disorder exemplified by X-linked adrenoleukodystrophy (ALD). Hundreds of ALD-causing mutations have been identified on ALDP. However, the pathogenic mechanisms of these mutations are restricted to clinical description due to limited structural and biochemical characterization. Here we report the cryo-electron microscopy structure of human ALDP with nominal resolution at 3.4 Å. ALDP exhibits a cytosolic-facing conformation. Compared to other lipid ATP-binding cassette transporters, ALDP has two substrate binding cavities formed by the transmembrane domains. Such structural organization may be suitable for the coordination of VLCFAs. Based on the structure, we performed integrative analysis of the cellular trafficking, protein thermostability, ATP hydrolysis, and the transport activity of representative mutations. These results provide a framework for understanding the working mechanism of ALDP and pathogenic roles of disease-associated mutations.
Topics: Humans; Adrenoleukodystrophy; Cryoelectron Microscopy; ATP Binding Cassette Transporter, Subfamily D, Member 1; ATP-Binding Cassette Transporters; Peroxisomes
PubMed: 36374178
DOI: 10.7554/eLife.75039 -
Analytical and Bioanalytical Chemistry Mar 2022Peroxisomes are versatile single membrane-enclosed cytoplasmic organelles, involved in reactive oxygen species (ROS) and lipid metabolism and diverse other metabolic...
Peroxisomes are versatile single membrane-enclosed cytoplasmic organelles, involved in reactive oxygen species (ROS) and lipid metabolism and diverse other metabolic processes. Peroxisomal disorders result from mutations in Pex genes-encoded proteins named peroxins (PEX proteins) and single peroxisomal enzyme deficiencies. The PEX11 protein family (α, β, and γ isoforms) plays an important role in peroxisomal proliferation and fission. However, their specific functions and the metabolic impact caused by their deficiencies have not been precisely characterized. To understand the systemic molecular alterations caused by peroxisomal defects, here we utilized untreated peroxisomal biogenesis factor 11α knockout (Pex11α KO) mouse model and performed serial relative-quantitative lipidomic, metabolomic, and proteomic analyses of serum, liver, and heart tissue homogenates. We demonstrated significant specific changes in the abundances of multiple lipid species, polar metabolites, and proteins and dysregulated metabolic pathways in distinct biological specimens of the Pex11α KO adult mice in comparison to the wild type (WT) controls. Overall, the present study reports comprehensive semi-quantitative molecular omics information of the Pex11α KO mice, which might serve in the future as a reference for a better understanding of the roles of Pex11α and underlying pathophysiological mechanisms of peroxisomal biogenesis disorders.
Topics: Animals; Lipidomics; Liver; Membrane Proteins; Mice; Mice, Knockout; Proteomics
PubMed: 35083512
DOI: 10.1007/s00216-021-03860-0 -
Biochemical Society Transactions Dec 2022Peroxisomes are membrane-bounded organelles that exist in most eukaryotic cells and are involved in the oxidation of fatty acids and the destruction of reactive oxygen...
Peroxisomes are membrane-bounded organelles that exist in most eukaryotic cells and are involved in the oxidation of fatty acids and the destruction of reactive oxygen species. Depending on the organism, they house additional metabolic reactions that range from glycolysis in parasitic protozoa to the production of ether lipids in animals and antibiotics in fungi. The importance of peroxisomes for human health is revealed by various disorders - notably the Zellweger spectrum - that are caused by defects in peroxisome biogenesis and are often fatal. Most peroxisomal metabolic enzymes reside in the lumen, but are synthesized in the cytosol and imported into the organelle by mobile receptors. The receptors accompany cargo all the way into the lumen and must return to the cytosol to start a new import cycle. Recycling requires receptor monoubiquitination by a membrane-embedded ubiquitin ligase complex composed of three RING finger (RF) domain-containing proteins: PEX2, PEX10, and PEX12. A recent cryo-electron microscopy (cryo-EM) structure of the complex reveals its function as a retro-translocation channel for peroxisomal import receptors. Each subunit of the complex contributes five transmembrane segments that assemble into an open channel. The N terminus of a receptor likely inserts into the pore from the lumenal side, and is then monoubiquitinated by one of the RFs to enable extraction into the cytosol. If recycling is compromised, receptors are polyubiquitinated by the concerted action of the other two RFs and ultimately degraded. The new data provide mechanistic insight into a crucial step of peroxisomal protein import.
Topics: Animals; Humans; Peroxins; Peroxisome-Targeting Signal 1 Receptor; Cryoelectron Microscopy; Membrane Proteins; Receptors, Cytoplasmic and Nuclear; Peroxisomes; Protein Transport; Ubiquitins; Ligases
PubMed: 36421406
DOI: 10.1042/BST20221393 -
Biomolecules Jul 2021Premature termination codon (PTC) mutations account for approximately 10% of pathogenic variants in monogenic diseases. Stimulation of translational readthrough, also...
Premature termination codon (PTC) mutations account for approximately 10% of pathogenic variants in monogenic diseases. Stimulation of translational readthrough, also known as stop codon suppression, using translational readthrough-inducing drugs (TRIDs) may serve as a possible therapeutic strategy for the treatment of genetic PTC diseases. One important parameter governing readthrough is the stop codon context (SCC)-the stop codon itself and the nucleotides in the vicinity of the stop codon on the mRNA. However, the quantitative influence of the SCC on treatment outcome and on appropriate drug concentrations are largely unknown. Here, we analyze the readthrough-stimulatory effect of various readthrough-inducing drugs on the SCCs of five common premature termination codon mutations of in a sensitive dual reporter system. Mutations in , encoding the peroxisomal targeting signal 1 receptor, can cause peroxisomal biogenesis disorders of the Zellweger spectrum. We show that the stop context has a strong influence on the levels of readthrough stimulation and impacts the choice of the most effective drug and its concentration. These results highlight potential advantages and the personalized medicine nature of an SCC-based strategy in the therapy of rare diseases.
Topics: Codon, Nonsense; HeLa Cells; Humans; Peroxisomal Disorders; Peroxisome-Targeting Signal 1 Receptor; Protein Biosynthesis; RNA, Messenger
PubMed: 34356630
DOI: 10.3390/biom11071006 -
Journal of Child Neurology Apr 2022Cerebral X-linked adrenoleukodystrophy (cALD) is an inflammatory demyelination of the brain that can lead to death unless treated by hematopoietic stem cell...
OBJECTIVES
Cerebral X-linked adrenoleukodystrophy (cALD) is an inflammatory demyelination of the brain that can lead to death unless treated by hematopoietic stem cell transplantation. Survival and improved outcomes for cerebral adrenoleukodystrophy are associated with hematopoietic stem cell transplantation at earliest evidence of disease on magnetic resonance imaging (MRI). Our goal was to determine average duration between diagnosis of cALD and hematopoietic stem cell transplantation.
METHODS
This was a retrospective review of data of patients aged 18 years or younger, using a nationwide administrative health care database (Pediatric Health Information System), with an () diagnosis of adrenoleukodystrophy. Time range was October 1, 2015, through June 30, 2021. We determined time to hematopoietic stem cell transplantation by duration between index brain MRI and a code for hematopoietic stem cell transplantation.
RESULTS
We identified 27 patients with cerebral adrenoleukodystrophy. Total charges for the cohort was $53 million. Time to transplant averaged 97 days. For Hispanic patients, time to transplant was 117 days, compared with 80 days for White, non-Hispanic patients. Comparison of different hospitals showed significant variability in time to hematopoietic stem cell transplantation.
DISCUSSION
We found that time to hematopoietic stem cell transplantation was >3 months for patients with cerebral adrenoleukodystrophy in the hospitals we evaluated. We noted differences in average time by race/ethnicity and by hospital. Our findings suggest opportunity to reduce time to transplant in cerebral adrenoleukodystrophy.
Topics: Adrenoleukodystrophy; Child; Hematopoietic Stem Cell Transplantation; Humans; Magnetic Resonance Imaging; Retrospective Studies
PubMed: 35238239
DOI: 10.1177/08830738221081141