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Proteome analysis of mouse cerebellar synaptosomes

ABSTRACT: In this study we have compared the proteomes from cerebellar synaptosome of early symptomatic Cstb-/- and wild type mice to gain insight into disease onset and progression of cystatin B deficiency. Biallelic loss-of-function mutations in the CSTB gene cause progressive myoclonus epilepsy of Unverricht-Lundborg type (EPM1). In the mouse model of EPM1, CSTB deficiency manifests from one month of age as progressive neurodegeneration, myoclonus and ataxia, which is preceded by neuroinflammation, alterations in GABAergic signaling, and a widespread rearrangement of the mitochondrial proteome in synapses.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Mus Musculus (mouse)

SUBMITTER: Tuula Nyman

LAB HEAD: Tuula Nyman

PROVIDER: PXD040382 | Pride | 2023-07-20

REPOSITORIES: pride

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Publications

Progressive mitochondrial dysfunction in cerebellar synaptosomes of cystatin B-deficient mice.

Gorski Katarin K Jackson Christopher B CB Nyman Tuula A TA Rezov Veronika V Battersby Brendan J BJ Lehesjoki Anna-Elina AE

Frontiers in molecular neuroscience 20230512

The involvement of mitochondrial dysfunction in cystatin B (CSTB) deficiency has been suggested, but its role in the onset of neurodegeneration, myoclonus, and ataxia in the CSTB-deficient mouse model (<i>Cstb<sup>-/-</sup></i>) is yet unknown. CSTB is an inhibitor of lysosomal and nuclear cysteine cathepsins. In humans, partial loss-of-function mutations cause the progressive myoclonus epilepsy neurodegenerative disorder, EPM1. Here we applied proteome analysis and respirometry on cerebellar sy ...[more]

PMID: 37251643

Similar Datasets

Proteomic analysis of cerebellar synaptosomes in presymptomatic cystatin B –deficient mice

Project description:Loss-of-function mutations in cystatin B (CSTB) cause progressive myoclonus epilepsy of Unverricht-Lundborg type (EPM1). Cstb-deficiency in mice leads to early alterations in GABAergic signaling, and causes neuroinflammation followed by progressive neurodegeneration, manifesting as progressive myoclonus and ataxia. The proteome of cerebellar synaptosomes of presymptomatic Cstb-/- mice were characterized by LC-ESI-MS/MS to gain insight into disease formation and progression.

2020-10-20 | PXD019370 | Pride

Gene expression alterations in the cerebellum and granule neurons of Cstb-/- mouse are associated with early synaptic changes and inflammation

Project description:Progressive myoclonus epilepsy of Unverricht-Lundborg type (EPM1) is an inherited neurodegenerative disease with myoclonus, seizures and ataxia, caused by the mutations in cystatin B (CSTB) gene. In an approach towards understanding the molecular basis of pathogenic events in EPM1 we have utilized the cystatin B deficient mice (Cstb-/-), a model for the disease. We have characterized the gene expression changes from the cerebellum of Cstb-/- mouse at postnatal day 7 (P7) and P30 as well as in cultured cerebellar granule cells using a pathway-based approach. A marked upregulation of immune response genes was seen at P30, reflecting the ongoing neuropathology, however, the observed alterations in complement cascade genes could also imply defects in synaptic plasticity. Differentially expressed genes in pre-symptomatic Cstb-/- animals at P7 were connected to synaptic function and plasticity and in cultured cerebellar granule cells to cellular biogenesis, cytoskeleton and intracellular transport. Especially GABAergic pathways were affected. The sample preparation and hybridzation of each cRNAs on GeneChipM-BM-. Mouse Genome 430 2.0 arrays (Affymetrix, Santa Clara, CA, USA) were performed in Helsinki Biomedicum Biochip Center (Finland).

2014-03-03 | E-GEOD-47516 | biostudies-arrayexpress

Gene expression alterations in the cerebellum and granule neurons of Cstb-/- mouse are associated with early synaptic changes and inflammation

2014-03-03 | GSE47516 | GEO

Gene expression data of cultured primary microglia from neonatal control and Cstb-/- mice.

Project description:Cystain B (Cstb) is a ubiquitously expressed cysteine protease inhibitor and mutations of the CSTB gene lead to the neurodegenerative disease progressive myoclonus epilepsy of Unverricht-Lundorg type (EPM1). We are interested in the microglia-specific, Cstb-dependent gene-expression changes in mice and in this data set, we include gene-level expression data from cultured primary microglia of control and Cstb-/- mice extracted from neonatal mice at P5 and we identified 156 differentially-expressed genes in Cstb-/- microglia.

2016-06-29 | GSE64823 | GEO

Gene expression data of cultured primary microglia from neonatal control and Cstb-/- mice.

2016-06-29 | E-GEOD-64823 | biostudies-arrayexpress

Progressive mitochondrial dysfunction in cerebellar synaptosomes of cystatin B-deficient mice.

Project description:The involvement of mitochondrial dysfunction in cystatin B (CSTB) deficiency has been suggested, but its role in the onset of neurodegeneration, myoclonus, and ataxia in the CSTB-deficient mouse model (Cstb-/-) is yet unknown. CSTB is an inhibitor of lysosomal and nuclear cysteine cathepsins. In humans, partial loss-of-function mutations cause the progressive myoclonus epilepsy neurodegenerative disorder, EPM1. Here we applied proteome analysis and respirometry on cerebellar synaptosomes from early symptomatic (Cstb-/-) mice to identify the molecular mechanisms involved in the onset of CSTB-deficiency associated neural pathogenesis. Proteome analysis showed that CSTB deficiency is associated with differential expression of mitochondrial and synaptic proteins, and respirometry revealed a progressive impairment in mitochondrial function coinciding with the onset of myoclonus and neurodegeneration in (Cstb-/-) mice. This mitochondrial dysfunction was not associated with alterations in mitochondrial DNA copy number or membrane ultrastructure. Collectively, our results show that CSTB deficiency generates a defect in synaptic mitochondrial bioenergetics that coincides with the onset and progression of the clinical phenotypes, and thus is likely a contributor to the pathogenesis of EPM1.

| S-EPMC10213208 | biostudies-literature

Quantitative Changes in the Mitochondrial Proteome of Cerebellar Synaptosomes From Preclinical Cystatin B-Deficient Mice.

Project description:Progressive myoclonus epilepsy of Unverricht-Lundborg type (EPM1) is a neurodegenerative disorder caused by loss-of-function mutations in the cystatin B (CSTB) gene. Progression of the clinical symptoms in EPM1 patients, including stimulus-sensitive myoclonus, tonic-clonic seizures, and ataxia, are well described. However, the cellular dysfunction during the presymptomatic phase that precedes the disease onset is not understood. CSTB deficiency leads to alterations in GABAergic signaling, and causes early neuroinflammation followed by progressive neurodegeneration in brains of a mouse model, manifesting as progressive myoclonus and ataxia. Here, we report the first proteome atlas from cerebellar synaptosomes of presymptomatic Cstb-deficient mice, and propose that early mitochondrial dysfunction is important to the pathogenesis of altered synaptic function in EPM1. A decreased sodium- and chloride dependent GABA transporter 1 (GAT-1) abundance was noted in synaptosomes with CSTB deficiency, but no functional difference was seen between the two genotypes in electrophysiological experiments with pharmacological block of GAT-1. Collectively, our findings provide novel insights into the early onset and pathogenesis of CSTB deficiency, and reveal greater complexity to the molecular pathogenesis of EPM1.

| S-EPMC7691638 | biostudies-literature

Pathological deficit of Cystatin B impairs synaptic plasticity in EPM1 human cerebral organoids

Project description:Cystatin B (CSTB) is a small protease inhibitor involved in cell proliferation and migration, and composition of extracellular matrix during brain development. Loss-of-function mutations in the gene encoding CSTB cause progressive myoclonic epilepsy 1 (EPM1). We previously demonstrated that CSTB is locally synthesized in the synaptosomes from rat brain and secreted into the media, indicating its role in synaptic plasticity. In this work, we further investigate the involvement of CSTB in synaptic plasticity, using synaptosomes from rodents’ brain, as well as from human Cerebral Organoids (hCOs). Our data demonstrated that CSTB is released by synaptosomes in two ways: as a soluble protein, and in extracellular vesicles-mediated manner. Synaptosomes isolated from hCOs are enriched in pre-synaptic proteins, as expected for an in vitro model of human synapses, and contain CSTB at all developmental stages analysed. However, only at late maturation stages synapse becomes physiologically active, i.e. particularly enriched with exocytosis-associated proteins. CSTB presence in the synaptic territories was confirmed also by immunostaining on human neurons in vitro. To investigate if the depletion of CSTB in EPM1 is affecting synaptic plasticity, we used synaptosomes from EPM1 hCOs. The synaptosomal levels of presynaptic proteins and of an initiation factor linked to local protein synthesis, were reduced in EPM1 hCOs compared to controls, and the extracellular vesicles trafficking was impaired. Moreover, morphological alteration was detected in neurons from EPM1 patients, indicating a connectivity deficit. In conclusion, our data indicate a significant role of synaptic CSTB in cell-to-cell communication in physiological and pathological conditions, and demonstrated that CSTB-dependent altered synaptic plasticity is one of the mechanisms underlying EPM1.

2024-08-10 | PXD041125 | Pride

Cystatin B is essential for proliferation and interneuron migration in individuals with EPM1 epilepsy

Project description:Progressive myoclonus epilepsy (PME) of Unverricht-Lundborg-type (EPM1) is an autosomal recessive neurodegenerative disorder with the highest incidence of PME worldwide. Mutations in the gene encoding cystatin B (CSTB) are the primary genetic cause of EPM1. Here, we investigate the role of CSTB during neurogenesis in vivo in the developing mouse brain and in vitro in human cerebral organoids (hCOs) derived from EPM1 patients. Remarkably, we find that CSTB (but not one of its pathological variants) is secreted into the mouse cerebral spinal fluid and the conditioned media from hCOs. In embryonic mouse brain, we find that functional CSTB influence progenitors’ proliferation and simultaneously modulate neuronal distribution by attracting interneurons to the site of secretion via cell non-autonomous mechanisms. Similarly, in patient-derived hCOs, low levels of functional CSTB result in an alteration of progenitor’s proliferation, premature differentiation, and changes in interneurons migration. Secretion and extracellular matrix organization are the biological processes particularly affected as revealed by proteomic analysis in patients’ hCOs. Overall, our data shed new light on the cellular mechanisms underlying the development of EPM1.

2020-05-13 | PXD018021 | Pride