Faithful SGCE imprinting in iPSC-derived cortical neurons: an endogenous cellular model of myoclonus-dystonia

In neuropathology research, induced pluripotent stem cell (iPSC)-derived neurons are considered a tool closely resembling the patient brain. Albeit in respect to epigenetics, this concept has been challenged. We generated iPSC-derived cortical neurons from myoclonus-dystonia patients with mutations (W100G and R102X) in the maternally imprinted ε-sarcoglycan (SGCE) gene and analysed properties such as imprinting, mRNA and protein expression. Comparison of the promoter during reprogramming and differentiation showed tissue-independent differential methylation. DNA sequencing with methylation-specific primers and cDNA analysis in patient neurons indicated selective expression of the mutated paternal SGCE allele. While fibroblasts only expressed the ubiquitous mRNA isoform, brain-specific SGCE mRNA and ε-sarcoglycan protein were detected in iPSC-derived control neurons. However, neuronal protein levels were reduced in both mutants. Our phenotypic characterization highlights the suitability of iPSC-derived cortical neurons with SGCE mutations for myoclonus-dystonia research and, in more general terms, prompts the use of iPSC-derived cellular models to study epigenetic mechanisms impacting on health and disease

Faithful SGCE imprinting in iPSC-derived cortical neurons: an endogenous cellular model of myoclonus-dystonia

In neuropathology research, induced pluripotent stem cell (iPSC)-derived neurons are considered a tool closely resembling the patient brain. Albeit in respect to epigenetics, this concept has been challenged. We generated iPSC-derived cortical neurons from myoclonus-dystonia patients with mutations (W100G and R102X) in the maternally imprinted ε-sarcoglycan (SGCE) gene and analysed properties such as imprinting, mRNA and protein expression. Comparison of the promoter during reprogramming and differentiation showed tissue-independent differential methylation. DNA sequencing with methylation-specific primers and cDNA analysis in patient neurons indicated selective expression of the mutated paternal SGCE allele. While fibroblasts only expressed the ubiquitous mRNA isoform, brain-specific SGCE mRNA and ε-sarcoglycan protein were detected in iPSC-derived control neurons. However, neuronal protein levels were reduced in both mutants. Our phenotypic characterization highlights the suitability of iPSC-derived cortical neurons with SGCE mutations for myoclonus-dystonia research and, in more general terms, prompts the use of iPSC-derived cellular models to study epigenetic mechanisms impacting on health and disease

Investigating the molecular and cellular basis of ε-sarcoglycan-associated myoclonus-dystonia in an iPSC-derived neuronal model

peer reviewedSGCE-related myoclonus-dystonia (M-D) underlies the epigenetic process of imprinting. This leads to the phenomenon of reduced penetrance upon maternal transmission of a pathogenic variant. Our previous work revealed that induced pluripotent stem cell (iPSC)-derived cortical neurons of carriers of pathogenic SGCE variants have been shown to serve as an adequate disease model for SGCE-related M-D, enabling the investigation of functional properties, such as the cellular localization of ε-sarcoglycan. Interestingly, ε-sarcoglycan has been linked to the dystrophin-associated glycoprotein complex (DGC) which is located at the plasma membrane and varies in its composition. Here, we investigate the molecular and cellular basis of epsilon-sarcoglycan-related myoclonus-dystonia in an iPSC-derived neuronal model from patients with pathogenic variants in SGCE.U-AGR-7238 - INTER/EJPRD22/17027921/PreDYT/Glaab (01/06/2023 - 31/05/2026) - GLAAB Enrico3. Good health and well-bein

Dystonia updates: definition, nomenclature, clinical classification, and etiology

A plethora of heterogeneous movement disorders is grouped under the umbrella term dystonia. The clinical presentation ranges from isolated dystonia to multi-systemic disorders where dystonia is only a co-occurring sign. In the past, definitions, nomenclature, and classifications have been repeatedly refined, adapted, and extended to reflect novel findings and increasing knowledge about the clinical, etiologic, and scientific background of dystonia. Currently, dystonia is suggested to be classified according to two axes. The first axis offers precise categories for the clinical presentation grouped into age at onset, body distribution, temporal pattern and associated features. The second, etiologic, axis discriminates pathological findings, as well as inheritance patterns, mode of acquisition, or unknown causality. Furthermore, the recent recommendations regarding terminology and nomenclature of inherited forms of dystonia and related syndromes are illustrated in this article. Harmonized, specific, and internationally widely used classifications provide the basis for future systematic dystonia research, as well as for more personalized patient counseling and treatment approaches

Impaired differentiation of human induced neural stem cells by TOR1A overexpression

DYT-TOR1A is the most common inherited dystonia caused by a three nucleotide (GAG) deletion (dE) in the TOR1A gene. Death early after birth and cortical anomalies of the full knockout in rodents underscore its developmental importance. We therefore explored the timed effects of TOR1A-wt and TOR1A-dE during differentiation in a human neural in vitro model. We used lentiviral tet-ON expression of TOR1A-wt and -dE in induced neural stem cells derived from healthy donors. Overexpression was induced during proliferation of neural precursors, during differentiation and after differentiation into mature neurons. Overexpression of both wildtype and mutated protein had no effect on the viability and cell number of neural precursors as well as mature neurons when initiated before or after differentiation. However, if induced during differentiation, overexpression of TOR1A-wt and -dE led to a pronounced reduction of mature neurons in a dose dependent manner. Our data underscores the importance of physiological expression levels of TOR1A as crucial for proper neuronal differentiation. We did not find evidence for a specific impact of the mutated TOR1A on neuronal maturation

Functional and Molecular Properties of DYT-SGCE Myoclonus-Dystonia Patient-Derived Striatal Medium Spiny Neurons.

Myoclonus-dystonia (DYT-SGCE, formerly DYT11) is characterized by alcohol-sensitive, myoclonic-like appearance of fast dystonic movements. It is caused by mutations in the SGCE gene encoding ε-sarcoglycan leading to a dysfunction of this transmembrane protein, alterations in the cerebello-thalamic pathway and impaired striatal plasticity. To elucidate underlying pathogenic mechanisms, we investigated induced pluripotent stem cell (iPSC)-derived striatal medium spiny neurons (MSNs) from two myoclonus-dystonia patients carrying a heterozygous mutation in the SGCE gene (c.298T>G and c.304C>T with protein changes W100G and R102X) in comparison to two matched healthy control lines. Calcium imaging showed significantly elevated basal intracellular Ca(2+) content and lower frequency of spontaneous Ca(2+) signals in SGCE MSNs. Blocking of voltage-gated Ca(2+) channels by verapamil was less efficient in suppressing KCl-induced Ca(2+) peaks of SGCE MSNs. Ca(2+) amplitudes upon glycine and acetylcholine applications were increased in SGCE MSNs, but not after GABA or glutamate applications. Expression of voltage-gated Ca(2+) channels and most ionotropic receptor subunits was not altered. SGCE MSNs showed significantly reduced GABAergic synaptic density. Whole-cell patch-clamp recordings displayed elevated amplitudes of miniature postsynaptic currents and action potentials in SGCE MSNs. Our data contribute to a better understanding of the pathophysiology and the development of novel therapeutic strategies for myoclonus-dystonia

Transcriptional Alterations in X-Linked Dystonia–Parkinsonism Caused by the SVA Retrotransposon

X-linked dystonia–parkinsonism (XDP) is a severe neurodegenerative disorder that manifests as adult-onset dystonia combined with parkinsonism. A SINE-VNTR-Alu (SVA) retrotransposon inserted in an intron of the TAF1 gene reduces its expression and alters splicing in XDP patient-derived cells. As a consequence, increased levels of the TAF1 intron retention transcript TAF1-32i can be found in XDP cells as compared to healthy controls. Here, we investigate the sequence of the deep intronic region included in this transcript and show that it is also present in cells from healthy individuals, albeit in lower amounts than in XDP cells, and that it undergoes degradation by nonsense-mediated mRNA decay. Furthermore, we investigate epigenetic marks (e.g., DNA methylation and histone modifications) present in this intronic region and the spanning sequence. Finally, we show that the SVA evinces regulatory potential, as demonstrated by its ability to repress the TAF1 promoter in vitro. Our results enable a better understanding of the disease mechanisms underlying XDP and transcriptional alterations caused by SVA retrotransposons