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1.
Acta Neuropathol ; 147(1): 67, 2024 04 06.
Article in English | MEDLINE | ID: mdl-38581586

ABSTRACT

Transcription factor EB (TFEB) is a master regulator of genes involved in the maintenance of autophagic and lysosomal homeostasis, processes which have been implicated in the pathogenesis of GBA-related and sporadic Parkinson's disease (PD), and dementia with Lewy bodies (DLB). TFEB activation results in its translocation from the cytosol to the nucleus. Here, we investigated TFEB subcellular localization and its relation to intracellular alpha-synuclein (aSyn) accumulation in post-mortem human brain of individuals with either incidental Lewy body disease (iLBD), GBA-related PD/DLB (GBA-PD/DLB) or sporadic PD/DLB (sPD/DLB), compared to control subjects. We analyzed nigral dopaminergic neurons using high-resolution confocal and stimulated emission depletion (STED) microscopy and semi-quantitatively scored the TFEB subcellular localization patterns. We observed reduced nuclear TFEB immunoreactivity in PD/DLB patients compared to controls, both in sporadic and GBA-related cases, as well as in iLBD cases. Nuclear depletion of TFEB was more pronounced in neurons with Ser129-phosphorylated (pSer129) aSyn accumulation in all groups. Importantly, we observed previously-unidentified TFEB-immunopositive perinuclear clusters in human dopaminergic neurons, which localized at the Golgi apparatus. These TFEB clusters were more frequently observed and more severe in iLBD, sPD/DLB and GBA-PD/DLB compared to controls, particularly in pSer129 aSyn-positive neurons, but also in neurons lacking detectable aSyn accumulation. In aSyn-negative cells, cytoplasmic TFEB clusters were more frequently observed in GBA-PD/DLB and iLBD patients, and correlated with reduced GBA enzymatic activity as well as increased Braak LB stage. Altered TFEB distribution was accompanied by a reduction in overall mRNA expression levels of selected TFEB-regulated genes, indicating a possible early dysfunction of lysosomal regulation. Overall, we observed cytoplasmic TFEB retention and accumulation at the Golgi in cells without apparent pSer129 aSyn accumulation in iLBD and PD/DLB patients. This suggests potential TFEB impairment at the early stages of cellular disease and underscores TFEB as a promising therapeutic target for synucleinopathies.


Subject(s)
Lewy Body Disease , Humans , alpha-Synuclein/metabolism , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/genetics , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/metabolism , Brain/pathology , Dopaminergic Neurons/metabolism , Lewy Bodies/pathology , Lewy Body Disease/pathology
2.
Nat Commun ; 14(1): 2057, 2023 04 12.
Article in English | MEDLINE | ID: mdl-37045813

ABSTRACT

Mutations in glucocerebrosidase cause the lysosomal storage disorder Gaucher's disease and are the most common risk factor for Parkinson's disease. Therapies to restore the enzyme's function in the brain hold great promise for treating the neurological implications. Thus, we developed blood-brain barrier penetrant therapeutic molecules by fusing transferrin receptor-binding moieties to ß-glucocerebrosidase (referred to as GCase-BS). We demonstrate that these fusion proteins show significantly increased uptake and lysosomal efficiency compared to the enzyme alone. In a cellular disease model, GCase-BS rapidly rescues the lysosomal proteome and lipid accumulations beyond known substrates. In a mouse disease model, intravenous injection of GCase-BS leads to a sustained reduction of glucosylsphingosine and can lower neurofilament-light chain plasma levels. Collectively, these findings demonstrate the potential of GCase-BS for treating GBA1-associated lysosomal dysfunction, provide insight into candidate biomarkers, and may ultimately open a promising treatment paradigm for lysosomal storage diseases extending beyond the central nervous system.


Subject(s)
Gaucher Disease , Parkinson Disease , Animals , Mice , Gaucher Disease/genetics , Glucosylceramidase/genetics , Glucosylceramidase/metabolism , Brain/metabolism , Neurons/metabolism , Parkinson Disease/metabolism , Lysosomes/metabolism , Mutation , alpha-Synuclein/metabolism
3.
Trends Neurosci ; 45(3): 184-199, 2022 03.
Article in English | MEDLINE | ID: mdl-35034773

ABSTRACT

The understanding of lysosomes has come a long way since the initial discovery of their role in degrading cellular waste. The lysosome is now recognized as a highly dynamic organelle positioned at the crossroads of cell signaling, transcription, and metabolism. Underscoring its importance is the observation that, in addition to rare monogenic lysosomal storage disorders, genes regulating lysosomal function are implicated in common sporadic neurodegenerative diseases, including Alzheimer's disease (AD), Parkinson's disease (PD), and amyotrophic lateral sclerosis (ALS). Developing therapies for these disorders is particularly challenging, largely due to gaps in knowledge of the underlying molecular and cellular processes. In this review, we discuss technological advances that have propelled deeper understanding of the lysosome in neurodegeneration, from elucidating the functions of lysosome-related disease risk variants at the level of the organelle, cell, and tissue, to the development of disease-specific biological models that recapitulate disease manifestations. Finally, we identify key questions to be addressed to successfully bridge the gap to the clinic.


Subject(s)
Alzheimer Disease , Amyotrophic Lateral Sclerosis , Parkinson Disease , Alzheimer Disease/metabolism , Amyotrophic Lateral Sclerosis/genetics , Humans , Lysosomes/metabolism , Parkinson Disease/metabolism
4.
Cell Rep ; 5(6): 1552-63, 2013 Dec 26.
Article in English | MEDLINE | ID: mdl-24373286

ABSTRACT

Abnormal accumulation of ß-secretase (BACE1) in dystrophic neurites and presynaptic ß-amyloid (Aß) production contribute to Alzheimer's disease pathogenesis. Little, however, is known about BACE1 sorting and dynamic transport in neurons. We investigated BACE1 trafficking in hippocampal neurons using live-cell imaging and selective labeling. We report that transport vesicles containing internalized BACE1 in dendrites undergo exclusive retrograde transport toward the soma, whereas they undergo bidirectional transport in axons. Unidirectional dendritic transport requires Eps15-homology-domain-containing (EHD) 1 and 3 protein function. Furthermore, loss of EHD function compromises dynamic axonal transport and overall BACE1 levels in axons. EHD1/3 colocalize with BACE1 and APP ß-C-terminal fragments in hippocampal mossy fiber terminals, and their depletion in neurons significantly attenuates Aß levels. These results demonstrate unidirectional endocytic transport of a dendritic cargo and reveal a role for EHD proteins in neuronal BACE1 transcytosis and Aß production, processes that are highly relevant for Alzheimer's disease.


Subject(s)
Alzheimer Disease/metabolism , Amyloid Precursor Protein Secretases/metabolism , Amyloid beta-Peptides/metabolism , Aspartic Acid Endopeptidases/metabolism , Axonal Transport , Carrier Proteins/metabolism , Dendrites/metabolism , Vesicular Transport Proteins/metabolism , Animals , Carrier Proteins/genetics , Cells, Cultured , HEK293 Cells , HeLa Cells , Hippocampus/cytology , Hippocampus/metabolism , Humans , Mice , Protein Transport , Vesicular Transport Proteins/genetics
5.
Cell Rep ; 5(6): 1536-51, 2013 Dec 26.
Article in English | MEDLINE | ID: mdl-24373285

ABSTRACT

Alzheimer's disease (AD) is characterized by cerebral deposition of ß-amyloid (Aß) peptides, which are generated from amyloid precursor protein (APP) by ß- and γ-secretases. APP and the secretases are membrane associated, but whether membrane trafficking controls Aß levels is unclear. Here, we performed an RNAi screen of all human Rab-GTPases, which regulate membrane trafficking, complemented with a Rab-GTPase-activating protein screen, and present a road map of the membrane-trafficking events regulating Aß production. We identify Rab11 and Rab3 as key players. Although retromers and retromer-associated proteins control APP recycling, we show that Rab11 controlled ß-secretase endosomal recycling to the plasma membrane and thus affected Aß production. Exome sequencing revealed a significant genetic association of Rab11A with late-onset AD, and network analysis identified Rab11A and Rab11B as components of the late-onset AD risk network, suggesting a causal link between Rab11 and AD. Our results reveal trafficking pathways that regulate Aß levels and show how systems biology approaches can unravel the molecular complexity underlying AD.


Subject(s)
Alzheimer Disease/metabolism , Amyloid beta-Peptides/metabolism , rab GTP-Binding Proteins/metabolism , Amyloid Precursor Protein Secretases/genetics , Amyloid Precursor Protein Secretases/metabolism , Aspartic Acid Endopeptidases/genetics , Aspartic Acid Endopeptidases/metabolism , Cell Membrane/metabolism , Endosomes/metabolism , Exome , GTPase-Activating Proteins/genetics , GTPase-Activating Proteins/metabolism , HeLa Cells , Humans , Protein Transport , Proteolysis , RNA, Small Interfering/genetics , rab GTP-Binding Proteins/genetics , rab3 GTP-Binding Proteins/genetics , rab3 GTP-Binding Proteins/metabolism
6.
ACS Chem Neurosci ; 4(7): 1057-61, 2013 Jul 17.
Article in English | MEDLINE | ID: mdl-23594172

ABSTRACT

Protein misfolding into amyloid-like aggregates underlies many neurodegenerative diseases. Thus, insights into the structure and function of these amyloids will provide valuable information on the pathological mechanisms involved and aid in the design of improved drugs for treating amyloid-based disorders. However, determining the structure of endogenous amyloids at high resolution has been difficult. Here we employ binding-activated localization microscopy (BALM) to acquire superresolution images of α-synuclein amyloid fibrils with unprecedented optical resolution. We propose that BALM imaging can be extended to study the structure of other amyloids, for differential diagnosis of amyloid-related diseases and for discovery of drugs that perturb amyloid structure for therapy.


Subject(s)
Amyloid/chemistry , Microscopy/methods , alpha-Synuclein/chemistry , Acetates , Humans , Thiophenes
7.
Trends Pharmacol Sci ; 33(4): 215-22, 2012 Apr.
Article in English | MEDLINE | ID: mdl-22385603

ABSTRACT

Bioavailability is a quantitative measure of the capacity of a drug to reach systemic circulation. However, if a drug target is localized in a subcellular organelle, then the drug may not be able to reach it and the effect of the drug will not be attained. Although most drug targets are localized within intracellular compartments, specific targeting of drugs at the subcellular level is not well established. Membrane proteins, lipids, nutrients and some pathogens are internalized into the cell to be targeted to distinct subcellular compartments via membrane trafficking. Recent advances have identified novel methods of subcellular drug targeting, involving the use of conjugation to ligands of cell surface receptors or to lipid anchors. In this review, we focus on the importance of subcellular targeting of drugs, in particular, the mechanism of lipid-anchoring as a novel strategy and its potential application for the treatment of several diseases.


Subject(s)
Intracellular Space/metabolism , Lipids/administration & dosage , Membrane Transport Proteins/metabolism , Pharmaceutical Preparations/administration & dosage , Receptors, Cell Surface/metabolism , Animals , Drug Delivery Systems/methods , Humans , Lipids/chemistry
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