Biology subjects

Lepine, S.

Publications and source records attributed to Lepine, S..

6 recordsLinked to original sources

bioRxiv2025

BetaII-Spectrin Gaps and Patches Emerge from the Patterned Assembly of the Actin/Spectrin Membrane Skeleton in Human Motor Neuron Axons

The actin/spectrin membrane-associated periodic skeleton (MPS) is a cytoskeletal structure that supports axonal integrity and function. Lower spinal motor neurons (MNs) are characterized by exceptionally long axons and are particularly susceptible to degeneration in a wide range of hereditary neuromuscular disorders, including amyotrophic lateral sclerosis. Using confocal and super-resolution imaging, we characterized the spatial distribution {beta}II-spectrin and the assembly pattern of the MPS in human MN axons derived from induced pluripotent stem cells (iPSCs). We discovered a striking gap-and-patch pattern in the medial axon, where sharply demarcated {beta}II-spectrin gaps alternate with patches containing a well-organized MPS. The pattern is acutely induced by the kinase inhibitor staurosporine and pharmacological inhibition of actin polymerization prevents patch formation, indicating a requirement for actin nucleation in MPS assembly. Our data supports a model in which spectrin incorporation into nascent MPS patches depletes neighboring regions, producing long-range gaps-and-patches patterns.

neuroscience↗

bioRxiv2025

Mutations in SOD1 induce ALS-related phenotypes in 3D iPSC-derived motor neuron (MN) spheroids

A significant challenge in ALS research is the heterogeneity of the disease. Even mutations within the same gene can lead to different disease prognosis. For instance, in silico protein modeling predicts distinct properties for distinct SOD1 mutations. With this in mind, in this study, we generated and characterized 3D iPSC-derived MN spheroids carrying homozygous knock-in SOD1 mutations (D90A and G93A), as well as a double mutation (D90A/G93A), to evaluate potential synergistic effects. An isogenic control line with the same genetic background was used for phenotypic comparisons with the knock-in variants. Mutant SOD1 MN spheroids exhibited multiple ALS-related phenotypes including altered SOD1 expression, reduced cell viability, downregulation of neurofilament (NF) subunit expression, hypoactivity, and altered burst activity. Our results highlight the advantages of using 3D MN spheroids as a disease model and stress the importance of considering phenotype variability at the genetic level in ALS.

neuroscience↗

bioRxiv2024

Transcriptome-based screening in TARDBP/TDP-43 knock-in motor neurons identifies the NEDD8-activating enzyme inhibitor MLN4924

A growing body of knowledge implicates perturbed RNA homeostasis in amyotrophic lateral sclerosis (ALS), a neurodegenerative disease that currently has no cure and few available treatments. Dysregulation of the multifunctional RNA-binding protein TDP-43 is increasingly regarded as a convergent feature of this disease, evidenced at the neuropathological level by the detection of TDP-43 pathology in most patient tissues, and at the genetic level by the identification of disease-associated mutations in its coding gene TARDBP. To characterize the transcriptional landscape induced by TARDBP mutations, we performed whole-transcriptome profiling of motor neurons differentiated from two knock-in iPSC lines expressing the ALS-linked TDP-43 variants p.A382T or p.G348C. Our results show that the TARDBP mutations significantly altered the expression profiles of mRNAs and microRNAs of the 14q32 cluster in MNs. Using mutation-induced gene signatures and the Connectivity Map database, we identified compounds predicted to restore gene expression toward wild-type levels. Among top-scoring compounds selected for further investigation, the NEDD8-activating enzyme inhibitor MLN4924 effectively improved cell viability and neuronal activity, highlighting a possible role for protein post-translational modification via NEDDylation in the pathobiology of TDP-43 in ALS.

neuroscience↗

bioRxiv2023

Early nuclear phenotypes and reactive transformation in human iPSC-derived astrocytes from ALS patients with SOD1 mutations

Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disease characterized by the progressive death of motor neurons (MNs). MN degeneration in ALS involves both cell-autonomous and non-cell autonomous mechanisms, with glial cells playing important roles in the latter. More specifically, astrocytes with mutations in the ALS-associated gene Cu/Zn superoxide dismutase 1 (SOD1) promote MN death. The mechanisms by which SOD1-mutated astrocytes reduce MN survival are incompletely understood. In order to characterize the impact of SOD1 mutations on astrocyte physiology, we generated astrocytes from human induced pluripotent stem cell (iPSC) derived from ALS patients carrying SOD1 mutations, together with control isogenic iPSCs. We report that astrocytes harbouring SOD1(A4V) and SOD1(D90A) mutations exhibit molecular and morphological changes indicative of reactive astrogliosis when compared to matching isogenic astrocytes. We show further that a number of nuclear phenotypes precede, or coincide with, reactive transformation. These include increased nuclear oxidative stress and DNA damage, and accumulation of the SOD1 protein in the nucleus. These findings reveal early cell-autonomous phenotypes in SOD1-mutated astrocytes that may contribute to the acquisition of a reactive phenotype involved in alterations of astrocyte-MN communication in ALS.

neuroscience↗

bioRxiv2023

Transcriptional dysregulation and impaired neuronal activity in FMR1 knock-out and Fragile X patients iPSC-derived models.

The lack of fragile X mental retardation protein (FMRP) protein, due to a repression of the FMR1 gene, causes Fragile X syndrome (FXS), one of the most prevalent forms of syndromic autisms. The FMR1 gene codes for an RNA binding protein involved in the regulation of gene expression through RNA processing, control of local translation, and protein-protein interactions; processes that are crucial for proper brain development. Taking advantage of induced pluripotent stem cells (iPSCs) and CRISPR-Cas9 genome editing technologies, we generated iPSC-derived cortical neural progenitors and cortical neurons from an FMR1 knock-out and patient cell line with the aim of identifying common phenotypes between the two cellular models. Using RNA sequencing, quantitative PCR and multielectrode array approaches, we assessed how the absence of the functional FMR1 gene affects the transcriptional profiles and the activities of iPSC-derived cortical neuronal progenitor cells (NPCs) and neurons with both models. We observed that FMR1 KO and FXS patient cells have a decrease in their mean firing rate; a cellular activity that can also be blocked by tetrodotoxin (TTX) application in wild-type active neurons. Relative to wild-type neurons, in FMR1 KO neurons, increased expression of presynaptic mRNA and transcription factors involved in the forebrain specification and decreased levels of mRNA coding AMPA and NMDA subunits were observed. Intriguingly, 40% of the differentially expressed genes were commonly deregulated between NPCs and differentiating neurons with significant enrichments in FMRP targets and Autism Related Genes found amongst downregulated genes. This implies that an absence of functional FMRP affects transcriptional profiles at the NPC stage, resulting in impaired activity and differentiation of the progenitors into mature neurons over time. These findings from the FMR1 KO lines were also shared with FXS patients iPSC-derived cells that also present with an impairment in activity and neuronal differentiation, illustrating the critical role of FMRP protein in neuronal development.

neuroscience↗

bioRxiv2023

Homozygous ALS-linked mutations in TARDBP/TDP-43 lead to progressive synaptic dysfunction in human iPSC-derived motor neurons

Cytoplasmic mislocalization and aggregation of the RNA-binding protein TDP-43 is a pathological hallmark of the motor neuron (MN) disease amyotrophic lateral sclerosis (ALS). Furthermore, while mutations in the TARDBP gene (encoding TDP-43) have been associated with ALS, the pathogenic consequences of these mutations remain poorly understood. Using CRISPR/Cas9, we engineered two homozygous knock-in iPSC lines carrying mutations in TARDBP encoding TDP-43A382T and TDP-43G348C, two common yet understudied ALS TDP-43 variants. MNs differentiated from knock-in iPSCs had normal viability and displayed no significant changes in TDP-43 subcellular localization, phosphorylation, solubility, or aggregation compared with isogenic control MNs. However, our results highlight synaptic impairments in both TDP-43A382T and TDP-43G348C MN cultures, as reflected in synapse abnormalities and alterations in spontaneous neuronal activity. Collectively, our findings suggest that MN dysfunction may precede the occurrence of TDP-43 pathology and neurodegeneration in ALS and further implicate synaptic and excitability defects in the pathobiology of this disease.

neuroscience↗