Publications by authors named "Patrik Verstreken"

115 Publications

MAPRE2 mutations result in altered human cranial neural crest migration, underlying craniofacial malformations in CSC-KT syndrome.

Sci Rep 2021 Mar 2;11(1):4976. Epub 2021 Mar 2.

Laboratory for the Genetics of Cognition, Department of Human Genetics, Center for Human Genetics, KU Leuven, Herestraat 49, 3000, Leuven, Belgium.

Circumferential skin creases (CSC-KT) is a rare polymalformative syndrome characterised by intellectual disability associated with skin creases on the limbs, and very characteristic craniofacial malformations. Previously, heterozygous and homozygous mutations in MAPRE2 were found to be causal for this disease. MAPRE2 encodes for a member of evolutionary conserved microtubule plus end tracking proteins, the end binding (EB) family. Unlike MAPRE1 and MAPRE3, MAPRE2 is not required for the persistent growth and stabilization of microtubules, but plays a role in other cellular processes such as mitotic progression and regulation of cell adhesion. The mutations identified in MAPRE2 all reside within the calponin homology domain, responsible to track and interact with the plus-end tip of growing microtubules, and previous data showed that altered dosage of MAPRE2 resulted in abnormal branchial arch patterning in zebrafish. In this study, we developed patient derived induced pluripotent stem cell lines for MAPRE2, together with isogenic controls, using CRISPR/Cas9 technology, and differentiated them towards neural crest cells with cranial identity. We show that changes in MAPRE2 lead to alterations in neural crest migration in vitro but also in vivo, following xenotransplantation of neural crest progenitors into developing chicken embryos. In addition, we provide evidence that changes in focal adhesion might underlie the altered cell motility of the MAPRE2 mutant cranial neural crest cells. Our data provide evidence that MAPRE2 is involved in cellular migration of cranial neural crest and offers critical insights into the mechanism underlying the craniofacial dysmorphisms and cleft palate present in CSC-KT patients. This adds the CSC-KT disorder to the growing list of neurocristopathies.
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http://dx.doi.org/10.1038/s41598-021-83771-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7925611PMC
March 2021

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition).

Autophagy 2021 Jan 8;17(1):1-382. Epub 2021 Feb 8.

University of Crete, School of Medicine, Laboratory of Clinical Microbiology and Microbial Pathogenesis, Voutes, Heraklion, Crete, Greece; Foundation for Research and Technology, Institute of Molecular Biology and Biotechnology (IMBB), Heraklion, Crete, Greece.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.
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http://dx.doi.org/10.1080/15548627.2020.1797280DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7996087PMC
January 2021

Lowering Synaptogyrin-3 expression rescues Tau-induced memory defects and synaptic loss in the presence of microglial activation.

Neuron 2021 03 19;109(5):767-777.e5. Epub 2021 Jan 19.

VIB-KU Leuven Center for Brain & Disease Research, Leuven, Belgium; KU Leuven, Department of Neurosciences, Leuven Brain Institute, Mission Lucidity, Leuven, Belgium. Electronic address:

Tau is a major driver of neurodegeneration and is implicated in over 20 diseases. Tauopathies are characterized by synaptic loss and neuroinflammation, but it is unclear if these pathological events are causally linked. Tau binds to Synaptogyrin-3 on synaptic vesicles. Here, we interfered with this function to determine the role of pathogenic Tau at pre-synaptic terminals. We show that heterozygous knockout of synaptogyrin-3 is benign in mice but strongly rescues mutant Tau-induced defects in long-term synaptic plasticity and working memory. It also significantly rescues the pre- and post-synaptic loss caused by mutant Tau. However, Tau-induced neuroinflammation remains clearly upregulated when we remove the expression of one allele of synaptogyrin-3. Hence neuroinflammation is not sufficient to cause synaptic loss, and these processes are separately induced in response to mutant Tau. In addition, the pre-synaptic defects caused by mutant Tau are enough to drive defects in cognitive tasks.
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http://dx.doi.org/10.1016/j.neuron.2020.12.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7927913PMC
March 2021

Maturation of neuronal AD-tau pathology involves site-specific phosphorylation of cytoplasmic and synaptic tau preceding conformational change and fibril formation.

Acta Neuropathol 2021 02 11;141(2):173-192. Epub 2021 Jan 11.

Laboratory for Neuropathology, Department of Imaging and Pathology, KU-Leuven, Herestraat 49, 3000, Leuven, Belgium.

In Alzheimer's disease (AD), tau-protein undergoes a multi-step process involving the transition from a natively unfolded monomer to large, aggregated structures such as neurofibrillary tangles (NFTs). However, it is not yet clear which events initiate the early preclinical phase of AD tauopathy and whether they have impact on the propagation of tau pathology in later disease stages. To address this question, we analyzed the distribution of tau species phosphorylated at T231, S396/S404 and S202/T205, conformationally modified at the MC1 epitope and fibrillary tau detected by the Gallyas method (Gallyas-tau), in the brains of 15 symptomatic and 20 asymptomatic cases with AD pathology as well as of 19 nonAD cases. As initial tau lesions, we identified phosphorylated-T231-tau diffusely distributed within the somatodendritic compartment (IC-tau) and phosphorylated-S396/pS404-tau in axonal lesions of the white matter and in the neuropil (IN-tau). The subcellular localization of pT231-tau in the cell body and pS396/pS404-tau in the presynapse was confirmed in hP301L mutant Drosophila larvae. Phosphorylated-S202/T205-tau, MC1-tau and Gallyas-tau were negative for these lesions. IC- and IN-tau were observed in all analyzed regions of the human brain, including early affected regions in nonAD cases (entorhinal cortex) and late affected regions in symptomatic AD cases (cerebellum), indicating that tau pathology initiation follows similar processes when propagating into previously unaffected regions. Furthermore, a sequence of AD-related maturation of tau-aggregates was observed, initiated by the appearance of IC- and IN-tau, followed by the formation of pretangles exhibiting pT231-tau, pS396/pS404-tau and pS202/pT205-tau, then by MC1-conformational tau, and, finally, by the formation of Gallyas-positive NFTs. Since cases classified as nonAD [Braak NFT stages < I (including a-1b)] already showed IC- and IN-tau, our findings suggest that these lesions are a prerequisite for the development of AD.
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http://dx.doi.org/10.1007/s00401-020-02251-6DOI Listing
February 2021

Molecule-to-Circuit Disease Mechanisms of a Synaptic SNAREopathy.

Neuron 2021 01;109(1):1-3

VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium; KU Leuven, Department of Neurosciences, Leuven Brain Institute, 3000 Leuven, Belgium. Electronic address:

Alten et al. present a detailed investigation of disease-causing SNAP25 mutations based on structural analysis, neurotransmitter release, and emerging circuit properties. They show that structurally clustered mutations within the SNAP25 SNARE motif cause similar functional defects and predict that alterations of spontaneous release are a novel disease mechanism.
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http://dx.doi.org/10.1016/j.neuron.2020.12.009DOI Listing
January 2021

A structure of substrate-bound Synaptojanin1 provides new insights in its mechanism and the effect of disease mutations.

Elife 2020 12 22;9. Epub 2020 Dec 22.

VIB-VUB Center for Structural Biology, Brussels, Belgium.

Synaptojanin1 (Synj1) is a phosphoinositide phosphatase, important in clathrin uncoating during endocytosis of presynaptic vesicles. It was identified as a potential drug target for Alzheimer's disease, Down syndrome, and TBC1D24-associated epilepsy, while also loss-of-function mutations in Synj1 are associated with epilepsy and Parkinson's disease. Despite its involvement in a range of disorders, structural, and detailed mechanistic information regarding the enzyme is lacking. Here, we report the crystal structure of the 5-phosphatase domain of Synj1. Moreover, we also present a structure of this domain bound to the substrate diC8-PI(3,4,5)P, providing the first image of a 5-phosphatase with a trapped substrate in its active site. Together with an analysis of the contribution of the different inositide phosphate groups to catalysis, these structures provide new insights in the Synj1 mechanism. Finally, we analysed the effect of three clinical missense mutations (Y793C, R800C, Y849C) on catalysis, unveiling the molecular mechanisms underlying Synj1-associated disease.
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http://dx.doi.org/10.7554/eLife.64922DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7781601PMC
December 2020

Excess Lipin enzyme activity contributes to TOR1A recessive disease and DYT-TOR1A dystonia.

Brain 2020 06;143(6):1746-1765

VIB-KU Leuven Center for Brain and Disease Research, Leuven, Belgium.

TOR1A/TorsinA mutations cause two incurable diseases: a recessive congenital syndrome that can be lethal, and a dominantly-inherited childhood-onset dystonia (DYT-TOR1A). TorsinA has been linked to phosphatidic acid lipid metabolism in Drosophila melanogaster. Here we evaluate the role of phosphatidic acid phosphatase (PAP) enzymes in TOR1A diseases using induced pluripotent stem cell-derived neurons from patients, and mouse models of recessive Tor1a disease. We find that Lipin PAP enzyme activity is abnormally elevated in human DYT-TOR1A dystonia patient cells and in the brains of four different Tor1a mouse models. Its severity also correlated with the dosage of Tor1a/TOR1A mutation. We assessed the role of excess Lipin activity in the neurological dysfunction of Tor1a disease mouse models by interbreeding these with Lpin1 knock-out mice. Genetic reduction of Lpin1 improved the survival of recessive Tor1a disease-model mice, alongside suppressing neurodegeneration, motor dysfunction, and nuclear membrane pathology. These data establish that TOR1A disease mutations cause abnormal phosphatidic acid metabolism, and suggest that approaches that suppress Lipin PAP enzyme activity could be therapeutically useful for TOR1A diseases.
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http://dx.doi.org/10.1093/brain/awaa139DOI Listing
June 2020

TBC1D24-TLDc-related epilepsy exercise-induced dystonia: rescue by antioxidants in a disease model.

Brain 2019 08;142(8):2319-2335

Pediatric Neurology, Neurogenetics and Neurobiology Unit and Laboratories, Children's Hospital A. Meyer-University of Florence, Florence, Italy.

Genetic mutations in TBC1D24 have been associated with multiple phenotypes, with epilepsy being the main clinical manifestation. The TBC1D24 protein consists of the unique association of a Tre2/Bub2/Cdc16 (TBC) domain and a TBC/lysin motif domain/catalytic (TLDc) domain. More than 50 missense and loss-of-function mutations have been described and are spread over the entire protein. Through whole genome/exome sequencing we identified compound heterozygous mutations, R360H and G501R, within the TLDc domain, in an index family with a Rolandic epilepsy exercise-induced dystonia phenotype (http://omim.org/entry/608105). A 20-year long clinical follow-up revealed that epilepsy was self-limited in all three affected patients, but exercise-induced dystonia persisted into adulthood in two. Furthermore, we identified three additional sporadic paediatric patients with a remarkably similar phenotype, two of whom had compound heterozygous mutations consisting of an in-frame deletion I81_K84 and an A500V mutation, and the third carried T182M and G511R missense mutations, overall revealing that all six patients harbour a missense mutation in the subdomain of TLDc between residues 500 and 511. We solved the crystal structure of the conserved Drosophila TLDc domain. This allowed us to predict destabilizing effects of the G501R and G511R mutations and, to a lesser degree, of R360H and potentially A500V. Next, we characterized the functional consequences of a strong and a weak TLDc mutation (TBC1D24G501R and TBC1D24R360H) using Drosophila, where TBC1D24/Skywalker regulates synaptic vesicle trafficking. In a Drosophila model neuronally expressing human TBC1D24, we demonstrated that the TBC1D24G501R TLDc mutation causes activity-induced locomotion and synaptic vesicle trafficking defects, while TBC1D24R360H is benign. The neuronal phenotypes of the TBC1D24G501R mutation are consistent with exacerbated oxidative stress sensitivity, which is rescued by treating TBC1D24G501R mutant animals with antioxidants N-acetylcysteine amide or α-tocopherol as indicated by restored synaptic vesicle trafficking levels and sustained behavioural activity. Our data thus show that mutations in the TLDc domain of TBC1D24 cause Rolandic-type focal motor epilepsy and exercise-induced dystonia. The humanized TBC1D24G501R fly model exhibits sustained activity and vesicle transport defects. We propose that the TBC1D24/Sky TLDc domain is a reactive oxygen species sensor mediating synaptic vesicle trafficking rates that, when dysfunctional, causes a movement disorder in patients and flies. The TLDc and TBC domain mutations' response to antioxidant treatment we observed in the animal model suggests a potential for combining antioxidant-based therapeutic approaches to TBC1D24-associated disorders with previously described lipid-altering strategies for TBC domain mutations.
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http://dx.doi.org/10.1093/brain/awz175DOI Listing
August 2019

Mitochondria Re-set Epilepsy.

Neuron 2019 06;102(5):907-910

VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium; KU Leuven, Department of Neurosciences, Leuven Brain Institute, 3000 Leuven, Belgium. Electronic address:

Neuronal networks maintain stable activity around a given set point, an enigmatic variable in homeostatic systems. In this issue of Neuron, Styr et al. (2019) now show that set points are regulated by mitochondria and propose a potential strategy to treat refractory forms of epilepsy.
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http://dx.doi.org/10.1016/j.neuron.2019.05.023DOI Listing
June 2019

Need for speed: Super-resolving the dynamic nanoclustering of syntaxin-1 at exocytic fusion sites.

Neuropharmacology 2020 06 28;169:107554. Epub 2019 Feb 28.

Clem Jones Centre for Ageing Dementia Research, Queensland Brain Institute, The University of Queensland, Queensland, Australia. Electronic address:

Communication between cells relies on regulated exocytosis, a multi-step process that involves the docking, priming and fusion of vesicles with the plasma membrane, culminating in the release of neurotransmitters and hormones. Key proteins and lipids involved in exocytosis are subjected to Brownian movement and constantly switch between distinct motion states which are governed by short-lived molecular interactions. Critical biochemical reactions between exocytic proteins that occur in the confinement of nanodomains underpin the precise sequence of priming steps which leads to the fusion of vesicles. The advent of super-resolution microscopy techniques has provided the means to visualize individual molecules on the plasma membrane with high spatiotemporal resolution in live cells. These techniques are revealing a highly dynamic nature of the nanoscale organization of the exocytic machinery. In this review, we focus on soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) syntaxin-1, which mediates vesicular fusion. Syntaxin-1 is highly mobile at the plasma membrane, and its inherent speed allows fast assembly and disassembly of syntaxin-1 nanoclusters which are associated with exocytosis. We reflect on recent studies which have revealed the mechanisms regulating syntaxin-1 nanoclustering on the plasma membrane and draw inferences on the effect of synaptic activity, phosphoinositides, N-ethylmaleimide-sensitive factor (NSF), α-soluble NSF attachment protein (α-SNAP) and SNARE complex assembly on the dynamic nanoscale organization of syntaxin-1. This article is part of the special issue entitled 'Mobility and trafficking of neuronal membrane proteins'.
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http://dx.doi.org/10.1016/j.neuropharm.2019.02.036DOI Listing
June 2020

Hsp90 Mediates Membrane Deformation and Exosome Release.

Mol Cell 2018 09;71(5):689-702.e9

VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium; KU Leuven, Department of Neurosciences, Leuven Brain Institute, 3000 Leuven, Belgium. Electronic address:

Hsp90 is an essential chaperone that guards proteome integrity and amounts to 2% of cellular protein. We now find that Hsp90 also has the ability to directly interact with and deform membranes via an evolutionarily conserved amphipathic helix. Using a new cell-free system and in vivo measurements, we show this amphipathic helix allows exosome release by promoting the fusion of multivesicular bodies (MVBs) with the plasma membrane. We dissect the relationship between Hsp90 conformation and membrane-deforming function and show that mutations and drugs that stabilize the open Hsp90 dimer expose the helix and allow MVB fusion, while these effects are blocked by the closed state. Hence, we structurally separated the Hsp90 membrane-deforming function from its well-characterized chaperone activity, and we show that this previously unrecognized function is required for exosome release.
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http://dx.doi.org/10.1016/j.molcel.2018.07.016DOI Listing
September 2018

Assaying Mutants of Clathrin-Mediated Endocytosis in the Fly Eye.

Methods Mol Biol 2018 ;1847:109-119

Laboratory of Neuronal Communication, KU Leuven, Department of Neurosciences, Leuven Brain Institute, VIB-KU Leuven Center for Brain & Disease Research, 49 Herestraat box 602, Leuven, 3000, Belgium.

Clathrin-mediated endocytosis plays essential roles both during and after development, and loss-of-function mutants affected in this process are mostly not viable. Different approaches have been developed to circumvent this limitation, including resorting to mosaic model organisms. We here describe the use of FLP/FRT-mediated mitotic recombination to generate Drosophila melanogaster having homozygous mutant eyes while the rest of their body is heterozygous. We then present a detailed protocol for assessing the consequences of these loss-of-function mutations on endocytosis in the photoreceptors of living fruit flies by recording electroretinograms.
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http://dx.doi.org/10.1007/978-1-4939-8719-1_9DOI Listing
April 2019

Parkinson's disease: convergence on synaptic homeostasis.

EMBO J 2018 09 31;37(18). Epub 2018 Jul 31.

VIB-KU Leuven Center for Brain& Disease Research, Leuven, Belgium

Parkinson's disease, the second most common neurodegenerative disorder, affects millions of people globally. There is no cure, and its prevalence will double by 2030. In recent years, numerous causative genes and risk factors for Parkinson's disease have been identified and more than half appear to function at the synapse. Subtle synaptic defects are thought to precede blunt neuronal death, but the mechanisms that are dysfunctional at synapses are only now being unraveled. Here, we review recent work and propose a model where different Parkinson proteins interact in a cell compartment-specific manner at the synapse where these proteins regulate endocytosis and autophagy. While this field is only recently emerging, the work suggests that the loss of synaptic homeostasis may contribute to neurodegeneration and is a key player in Parkinson's disease.
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http://dx.doi.org/10.15252/embj.201898960DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6138432PMC
September 2018

Imaging mitophagy in the fruit fly.

Autophagy 2018 2;14(9):1656-1657. Epub 2018 Aug 2.

a Laboratory for Parkinson Research, Department of Neurosciences , Leuven Brain Institute , Leuven , Belgium.

Loss-of-function mutations in the genes encoding PRKN/parkin and PINK1 cause autosomal recessive Parkinson disease (PD). Seminal work in Drosophila revealed that loss of park/parkin and Pink1 causes prominent mitochondrial pathology in flight muscle and, to a lesser extent, in dopaminergic neurons. Subsequent studies in cultured mammalian cells discovered a crucial role for PRKN/PARK2 and PINK1 in selective macroautophagic removal of mitochondria (mitophagy). However, direct evidence for the existence of a PINK1-PRKN/PARK2-mediated mitophagy pathway in vivo is still scarce. Recently, we engineered Drosophila that express the mitophagy reporter mt-Keima. We demonstrated that mitophagy occurs in flight muscle cells and dopaminergic neurons in vivo and increases with aging. Moreover, this age-dependent rise depends on park and Pink1. Our data also suggested that some aspects of the mitochondrial phenotype of park- and Pink1-deficient flies are independent of the mitophagy defect, and that park and Pink1 may have multiple functions in the regulation of the integrity of these organelles. Here, we discuss implications of these findings as well as possible future applications of the mt-Keima fly model.
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http://dx.doi.org/10.1080/15548627.2018.1496720DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6135567PMC
July 2019

A Single-Cell Transcriptome Atlas of the Aging Drosophila Brain.

Cell 2018 08 18;174(4):982-998.e20. Epub 2018 Jun 18.

VIB Center for Brain & Disease Research, KU Leuven, Leuven 3000, Belgium; Department of Human Genetics KU Leuven, Leuven 3000, Belgium. Electronic address:

The diversity of cell types and regulatory states in the brain, and how these change during aging, remains largely unknown. We present a single-cell transcriptome atlas of the entire adult Drosophila melanogaster brain sampled across its lifespan. Cell clustering identified 87 initial cell clusters that are further subclustered and validated by targeted cell-sorting. Our data show high granularity and identify a wide range of cell types. Gene network analyses using SCENIC revealed regulatory heterogeneity linked to energy consumption. During aging, RNA content declines exponentially without affecting neuronal identity in old brains. This single-cell brain atlas covers nearly all cells in the normal brain and provides the tools to study cellular diversity alongside other Drosophila and mammalian single-cell datasets in our unique single-cell analysis platform: SCope (http://scope.aertslab.org). These results, together with SCope, allow comprehensive exploration of all transcriptional states of an entire aging brain.
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http://dx.doi.org/10.1016/j.cell.2018.05.057DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6086935PMC
August 2018

ER Lipid Defects in Neuropeptidergic Neurons Impair Sleep Patterns in Parkinson's Disease.

Neuron 2018 06 7;98(6):1155-1169.e6. Epub 2018 Jun 7.

VIB-KU Leuven Center for Brain and Disease Research, 3000 Leuven, Belgium; Department of Neurosciences, Leuven Brain Institute, KU Leuven, 3000 Leuven, Belgium. Electronic address:

Parkinson's disease patients report disturbed sleep patterns long before motor dysfunction. Here, in parkin and pink1 models, we identify circadian rhythm and sleep pattern defects and map these to specific neuropeptidergic neurons in fly models and in hypothalamic neurons differentiated from patient induced pluripotent stem cells (iPSCs). Parkin and Pink1 control the clearance of mitochondria by protein ubiquitination. Although we do not observe major defects in mitochondria of mutant neuropeptidergic neurons, we do find an excess of endoplasmic reticulum-mitochondrial contacts. These excessive contact sites cause abnormal lipid trafficking that depletes phosphatidylserine from the endoplasmic reticulum (ER) and disrupts the production of neuropeptide-containing vesicles. Feeding mutant animals phosphatidylserine rescues neuropeptidergic vesicle production and acutely restores normal sleep patterns in mutant animals. Hence, sleep patterns and circadian disturbances in Parkinson's disease models are explained by excessive ER-mitochondrial contacts, and blocking their formation or increasing phosphatidylserine levels rescues the defects in vivo.
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http://dx.doi.org/10.1016/j.neuron.2018.05.022DOI Listing
June 2018

Deficiency of parkin and PINK1 impairs age-dependent mitophagy in .

Elife 2018 05 29;7. Epub 2018 May 29.

Laboratory for Parkinson Research, Department of Neurosciences, Leuven, Belgium.

Mutations in the genes for PINK1 and parkin cause Parkinson's disease. PINK1 and parkin cooperate in the selective autophagic degradation of damaged mitochondria (mitophagy) in cultured cells. However, evidence for their role in mitophagy in vivo is still scarce. Here, we generated a model expressing the mitophagy probe mt-Keima. Using live mt-Keima imaging and correlative light and electron microscopy (CLEM), we show that mitophagy occurs in muscle cells and dopaminergic neurons in vivo, even in the absence of exogenous mitochondrial toxins. Mitophagy increases with aging, and this age-dependent rise is abrogated by PINK1 or parkin deficiency. Knockdown of the homologues of the deubiquitinases USP15 and, to a lesser extent, USP30, rescues mitophagy in the parkin-deficient flies. These data demonstrate a crucial role for parkin and PINK1 in age-dependent mitophagy in in vivo.
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http://dx.doi.org/10.7554/eLife.35878DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6008047PMC
May 2018

p13 protects against Parkinson's disease.

EMBO Rep 2018 Mar 19;19(3). Epub 2018 Feb 19.

VIB-KU Leuven Center for Brain & Disease Research, Leuven, Belgium.

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http://dx.doi.org/10.15252/embr.201845900DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5836093PMC
March 2018

Synaptogyrin-3 Mediates Presynaptic Dysfunction Induced by Tau.

Neuron 2018 02 1;97(4):823-835.e8. Epub 2018 Feb 1.

VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium; Department of Neurosciences and Leuven Brain Institute, KU Leuven, 3000 Leuven, Belgium. Electronic address:

Synaptic dysfunction is an early pathological feature of neurodegenerative diseases associated with Tau, including Alzheimer's disease. Interfering with early synaptic dysfunction may be therapeutically beneficial to prevent cognitive decline and disease progression, but the mechanisms underlying synaptic defects associated with Tau are unclear. In disease conditions, Tau mislocalizes into pre- and postsynaptic compartments; here we show that, under pathological conditions, Tau binds to presynaptic vesicles in Alzheimer's disease patient brain. We define that the binding of Tau to synaptic vesicles is mediated by the transmembrane vesicle protein Synaptogyrin-3. In fly and mouse models of Tauopathy, reduction of Synaptogyrin-3 prevents the association of presynaptic Tau with vesicles, alleviates Tau-induced defects in vesicle mobility, and restores neurotransmitter release. This work therefore identifies Synaptogyrin-3 as the binding partner of Tau on synaptic vesicles, revealing a new presynapse-specific Tau interactor, which may contribute to early synaptic dysfunction in neurodegenerative diseases associated with Tau.
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http://dx.doi.org/10.1016/j.neuron.2018.01.022DOI Listing
February 2018

In Vivo Single-Molecule Tracking at the Drosophila Presynaptic Motor Nerve Terminal.

J Vis Exp 2018 01 14(131). Epub 2018 Jan 14.

Clem Jones Centre for Ageing Dementia Research, Queensland Brain Institute, The University of Queensland;

An increasing number of super-resolution microscopy techniques are helping to uncover the mechanisms that govern the nanoscale cellular world. Single-molecule imaging is gaining momentum as it provides exceptional access to the visualization of individual molecules in living cells. Here, we describe a technique that we developed to perform single-particle tracking photo-activated localization microscopy (sptPALM) in Drosophila larvae. Synaptic communication relies on key presynaptic proteins that act by docking, priming, and promoting the fusion of neurotransmitter-containing vesicles with the plasma membrane. A range of protein-protein and protein-lipid interactions tightly regulates these processes and the presynaptic proteins therefore exhibit changes in mobility associated with each of these key events. Investigating how mobility of these proteins correlates with their physiological function in an intact live animal is essential to understanding their precise mechanism of action. Extracting protein mobility with high resolution in vivo requires overcoming limitations such as optical transparency, accessibility, and penetration depth. We describe how photoconvertible fluorescent proteins tagged to the presynaptic protein Syntaxin-1A can be visualized via slight oblique illumination and tracked at the motor nerve terminal or along the motor neuron axon of the third instar Drosophila larva.
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http://dx.doi.org/10.3791/56952DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5908646PMC
January 2018

Reprogramming neurodegeneration in the big data era.

Curr Opin Neurobiol 2018 02 10;48:167-173. Epub 2018 Jan 10.

VIB-KU Leuven Center for Brain & Disease Research, Leuven 3000, Belgium; Department of Neurosciences, Leuven Brain Institute, KU Leuven, 3000 Leuven, Belgium. Electronic address:

Recent genome-wide association studies (GWAS) have identified numerous genetic risk variants for late-onset Alzheimer's disease (AD) and Parkinson's disease (PD). However, deciphering the functional consequences of GWAS data is challenging due to a lack of reliable model systems to study the genetic variants that are often of low penetrance and non-coding identities. Pluripotent stem cell (PSC) technologies offer unprecedented opportunities for molecular phenotyping of GWAS variants in human neurons and microglia. Moreover, rapid technological advances in whole-genome RNA-sequencing and epigenome mapping fuel comprehensive and unbiased investigations of molecular alterations in PSC-derived disease models. Here, we review and discuss how integrated studies that utilize PSC technologies and genome-wide approaches may bring new mechanistic insight into the pathogenesis of AD and PD.
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http://dx.doi.org/10.1016/j.conb.2017.12.015DOI Listing
February 2018

Trapping of Syntaxin1a in Presynaptic Nanoclusters by a Clinically Relevant General Anesthetic.

Cell Rep 2018 01;22(2):427-440

Queensland Brain Institute, The University of Queensland, Brisbane QLD 4072, Australia. Electronic address:

Propofol is the most commonly used general anesthetic in humans. Our understanding of its mechanism of action has focused on its capacity to potentiate inhibitory systems in the brain. However, it is unknown whether other neural mechanisms are involved in general anesthesia. Here, we demonstrate that the synaptic release machinery is also a target. Using single-particle tracking photoactivation localization microscopy, we show that clinically relevant concentrations of propofol and etomidate restrict syntaxin1A mobility on the plasma membrane, whereas non-anesthetic analogs produce the opposite effect and increase syntaxin1A mobility. Removing the interaction with the t-SNARE partner SNAP-25 abolishes propofol-induced syntaxin1A confinement, indicating that syntaxin1A and SNAP-25 together form an emergent drug target. Impaired syntaxin1A mobility and exocytosis under propofol are both rescued by co-expressing a truncated syntaxin1A construct that interacts with SNAP-25. Our results suggest that propofol interferes with a step in SNARE complex formation, resulting in non-functional syntaxin1A nanoclusters.
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http://dx.doi.org/10.1016/j.celrep.2017.12.054DOI Listing
January 2018

α-Synuclein and Tau: Mitochondrial Kill Switches.

Neuron 2018 01;97(1):3-4

VIB-KU Leuven Center for Brain and Disease Research, 3000 Leuven, Belgium; KU Leuven, Department of Neurosciences, Leuven Brain Institute, 3000 Leuven, Belgium. Electronic address:

α-Synuclein resides in Lewy bodies in Parkinson's disease. Ordonez et al. (2017) now show that α-syn disrupts the actin network, causing Drp1-dependent mitochondrial fission defects. This is similar to defects induced by the PD risk factor Tau, suggesting converging pathways in neurodegeneration.
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http://dx.doi.org/10.1016/j.neuron.2017.12.024DOI Listing
January 2018

Autophagy in the presynaptic compartment in health and disease.

J Cell Biol 2017 07 17;216(7):1895-1906. Epub 2017 May 17.

Department of Neurosciences, Katholieke University Leuven, 3000 Leuven, Belgium.

Synapses are functionally distinct neuronal compartments that are critical for brain function, with synaptic dysfunction being an early pathological feature in aging and disease. Given the large number of proteins needed for synaptic function, the proliferation of defective proteins and the subsequent loss of protein homeostasis may be a leading cause of synaptic dysfunction. Autophagic mechanisms are cellular digestion processes that recycle cellular components and contribute to protein homeostasis. Autophagy is important within the nervous system, but its function in specific compartments such as the synapse has been unclear. Evidence from research on both autophagy and synaptic function suggests that there are links between the two and that synaptic homeostasis during aging requires autophagy to regulate protein homeostasis. Exciting new work on autophagy-modulating proteins that are enriched at the synapse has begun to link autophagy to synapses and synaptic dysfunction in disease. A better understanding of these links will help us harness the potential therapeutic benefits of autophagy in combating age-related disorders of the nervous system.
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http://dx.doi.org/10.1083/jcb.201611113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5496617PMC
July 2017

Tau association with synaptic vesicles causes presynaptic dysfunction.

Nat Commun 2017 05 11;8:15295. Epub 2017 May 11.

VIB-KU Leuven Center for Brain &Disease Research, Leuven 3000, Belgium.

Tau is implicated in more than 20 neurodegenerative diseases, including Alzheimer's disease. Under pathological conditions, Tau dissociates from axonal microtubules and missorts to pre- and postsynaptic terminals. Patients suffer from early synaptic dysfunction prior to Tau aggregate formation, but the underlying mechanism is unclear. Here we show that pathogenic Tau binds to synaptic vesicles via its N-terminal domain and interferes with presynaptic functions, including synaptic vesicle mobility and release rate, lowering neurotransmission in fly and rat neurons. Pathological Tau mutants lacking the vesicle binding domain still localize to the presynaptic compartment but do not impair synaptic function in fly neurons. Moreover, an exogenously applied membrane-permeable peptide that competes for Tau-vesicle binding suppresses Tau-induced synaptic toxicity in rat neurons. Our work uncovers a presynaptic role of Tau that may be part of the early pathology in various Tauopathies and could be exploited therapeutically.
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http://dx.doi.org/10.1038/ncomms15295DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5437271PMC
May 2017

The SAC1 domain in synaptojanin is required for autophagosome maturation at presynaptic terminals.

EMBO J 2017 05 22;36(10):1392-1411. Epub 2017 Mar 22.

VIB Center for Brain & Disease Research, Leuven, Belgium

Presynaptic terminals are metabolically active and accrue damage through continuous vesicle cycling. How synapses locally regulate protein homeostasis is poorly understood. We show that the presynaptic lipid phosphatase synaptojanin is required for macroautophagy, and this role is inhibited by the Parkinson's disease mutation R258Q. Synaptojanin drives synaptic endocytosis by dephosphorylating PI(4,5)P, but this function appears normal in knock-in flies. Instead, R258Q affects the synaptojanin SAC1 domain that dephosphorylates PI(3)P and PI(3,5)P, two lipids found in autophagosomal membranes. Using advanced imaging, we show that mutants accumulate the PI(3)P/PI(3,5)P-binding protein Atg18a on nascent synaptic autophagosomes, blocking autophagosome maturation at fly synapses and in neurites of human patient induced pluripotent stem cell-derived neurons. Additionally, we observe neurodegeneration, including dopaminergic neuron loss, in flies. Thus, synaptojanin is essential for macroautophagy within presynaptic terminals, coupling protein turnover with synaptic vesicle cycling and linking presynaptic-specific autophagy defects to Parkinson's disease.
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http://dx.doi.org/10.15252/embj.201695773DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5430236PMC
May 2017

Neurons eat glutamate to stay alive.

J Cell Biol 2017 04 15;216(4):863-865. Epub 2017 Mar 15.

VIB Center for Brain and Disease Research, Katholieke Universiteit Leuven, 3000 Leuven, Belgium

Neurons are thought to primarily rely on glucose to fuel mitochondrial metabolism. In this issue, Divakaruni et al. (2017. https://doi.org/10.1083/jcb.201612067) show that neurons are also happy to use glutamate. When neurons use this neurotransmitter, its concentration drops, thus protecting against glutamate-induced excitotoxic stress.
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http://dx.doi.org/10.1083/jcb.201702003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5379959PMC
April 2017

EndoA/Endophilin-A creates docking stations for autophagic proteins at synapses.

Autophagy 2017 May 15;13(5):971-972. Epub 2017 Feb 15.

a VIB Center for Brain & Disease Research , Leuven , Belgium.

Synapses are very specialized compartments with high metabolic demand to maintain neurotransmission, an essential step for basic brain function. Neurons are post-mitotic and synapses need to stay functional over time-sometimes over decades. Given that synapses are often at a long distance from the cell body, they must use local mechanisms to regulate protein quality control. We show that macroautophagy/autophagy is one of these local processes and found that it is under strict control of the synapse-enriched protein EndoA/Endophilin-A, previously only implicated in endocytosis. Metabolic and neuronal stimulation induce synaptic autophagy and phosphorylation of EndoA by the Parkinson disease kinase Lrrk/LRRK2 is essential to promote the process. EndoA induces membrane curvature in vitro, and, mechanistically, phosphorylated EndoA creates curved membrane-protein docking sites that are capable of recruiting Atg3. Our work reveals a synapse-enriched branch of autophagy under the control of EndoA that may be deregulated in Parkinson disease.
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http://dx.doi.org/10.1080/15548627.2017.1286440DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5446079PMC
May 2017

Presynaptic protein homeostasis and neuronal function.

Curr Opin Genet Dev 2017 Jun 16;44:38-46. Epub 2017 Feb 16.

KU Leuven, Department of Neurosciences, Leuven Institute for Neurodegenerative Disease (LIND), Herestraat 49, bus 602, 3000 Leuven, Belgium; VIB Center for Brain & Disease Research, Herestraat 49, bus 602, 3000 Leuven, Belgium. Electronic address:

Proteome integrity is maintained by a coordinated network of molecular chaperones, by protein degradation machineries and by their regulators. Numerous human pathologies are considered as diseases of compromised protein homeostasis (proteostasis), including neurodegeneration. These are characterized by the accumulation of neuronal protein aggregates and by synaptic defects followed by loss of connectivity and cell death. While this suggests that synaptic terminals are particularly sensitive to proteostasis imbalance, our understanding of protein turnover mechanisms and regulation at the synapse remains limited. Recent reports show that different proteolytic pathways act at synapses, including several forms of autophagy. The role of chaperones in controlling the balance between synaptic protein refolding and degradation and how this complex network regulates neuronal function also begins to be unraveled.
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http://dx.doi.org/10.1016/j.gde.2017.01.015DOI Listing
June 2017