Biology subjects

Kabashi, E.

Publications and source records attributed to Kabashi, E..

7 recordsLinked to original sources

bioRxiv2025

Multiplex imaging combined to machine learning enable automated profiling of cortical malformations applications in tuberous sclerosis complex

Malformations of cortical development such as tuberous sclerosis complex arise within a heterogeneous cellular landscape that conventional histopathology only partially resolves. Here, we combined a 19-marker multiplex immunofluorescence panel with a machine learning-driven image analysis pipeline to map and quantify over 365 000 cells from paediatric surgical cortex, defining the single-cell architecture of TSC lesions. Microtubers were objectively delineated by vimentin and detected in all TSC samples but absent from non-dysplastic controls. Within these structures, balloon cells exhibiting strong pS6 activation occupied lesion cores and were confined to microtuber boundaries, whereas dysmorphic neurons were more diffusely distributed into adjacent cortex. The microtuber niche was dominated by astroglial remodeling: immature and reactive vimentin-positive astrocytes, including Lamp5-positive subsets, accumulated at and around lesion rims, while mature GFAP-positive astrocytes showed only modest changes. Distance-based spatial analyses revealed neuronal exclusion from microtuber centres with gradual recovery in surrounding tissue, indicating local network disruption. Unsupervised clustering and niche modelling recapitulated these spatial gradients, identifying a glial-dominated ecosystem that concentrates balloon cells, increases inter-neuronal distances, and reduces cell-cell interactions. Together, these data support a model in which cortical tubers arise through the coalescence of microtubers orchestrated by balloon cells and reactive gliosis during corticogenesis. Beyond elucidating disease architecture, our automated framework enables reproducible lesion detection, quantitative cell-state mapping, and spatial readouts applicable across malformations of cortical development. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=126 SRC="FIGDIR/small/690101v1_ufig1.gif" ALT="Figure 1"> View larger version (57K): org.highwire.dtl.DTLVardef@dc3258org.highwire.dtl.DTLVardef@6d4149org.highwire.dtl.DTLVardef@d93354org.highwire.dtl.DTLVardef@1b8f83d_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Integrative Analysis of Drug-Gene Expression Signatures in Human Pluripotent Stem Cells Identifies Novel Drug Candidates for ALS and Monogenic Diseases

The classical paradigm of drug screening often faces significant limitations due to the challenges associated with identifying molecular or cellular read-outs that are relevant to specific diseases. This issue is particularly pronounced for the thousands of diseases of genetic origin, given the abundance of databases listing disease-associated changes in gene expression and various transcripts, indicating that potential read-outs may be concealed within these resources. To remedy this, an alternative approach was tested: compounds were evaluated for their effects on gene expression and alternative splicing in a healthy cell model, and the resulting data were matched to molecular signatures of diseases. A subset of 50 FDA-approved drugs was tested on mesenchymal stem cells derived from a human pluripotent stem cell line. Over half of the compounds altered gene expression, many affecting pathways linked to monogenic diseases. One hit, increased SQSTM1 expression induced by prazosin, was further validated in ALS models caused by SQSTM1 haploinsufficiency, including patient-derived fibroblasts, SQSTM1-depleted hiPSC-derived motor neurons, and a zebrafish model. Extending this paradigm could involve testing diverse cell types and larger drug libraries. One Sentence SummaryA novel drug screening approach for monogenic diseases integrating human pluripotent stem cell derivatives and RNA sequencing to profile gene expression.

genomics↗

bioRxiv2024

Alternative splicing of SORBS1 affects neuromuscular junction formation and stability in myotonic dystrophy type 1

Myotonic dystrophy type 1 (DM1) is a multisystemic neuromuscular disease characterized by a CTG repeat expansion in the 3" untranslated region of the gene coding for the dystrophia myotonica protein kinase (DMPK). Presence of expanded CTG repeats in DMPK-mRNAs leads to the sequestration of RNA binding factors such as the Muscleblind like (MBNL) proteins resulting in widespread splicing defects contributing to progressive muscle weakness and myotonia. Here, we show that abnormal splicing of SORBS1 exon 25 found in skeletal muscle of myotonic dystrophy type 1 patients is a critical contributor to neuromuscular junction (NMJ) formation and maintenance. Forced exclusion of SORBS1 exon 25 in mice results in NMJ degeneration with marked denervation and postsynaptic destabilization. In zebrafish, misregulation of sorbs1 exon 25 results in reduced motor function and abnormal AChR cluster morphology. Finally, we observed that forcing SORBS1 exon 25 exclusion in hiPSC-derived skeletal muscle cells reduces the formation of large AChR clusters upon agrin stimulation. Thus, our study identifies MBNL regulated SORBS1 alternative splicing during skeletal muscle development as a critical event for NMJ formation and maintenance. The aberrant splicing of SORBS1 exon 25 in DM1 expands our understanding of how splicing dysregulation compromises neuromuscular system communication, shedding light on the broader impact of mRNA splicing regulation on NMJ integrity.

molecular biology↗

bioRxiv2024

Functional assessment of a kcnb1 knock-out zebrafish to model KCNB1-related neurodevelopmental and epileptic disorders

KEY POINTSO_LIkcnb1 is expressed in distinct cell subtypes and various regions of the central nervous system in zebrafish C_LIO_LIBrain anatomy and neuronal circuits are not disrupted in the kcnb1 loss-of-function zebrafish model C_LIO_LILoss of kcnb1 leads to altered behavior phenotype, light and sound-induced locomotor impairments C_LIO_LIkcnb1 knock-out zebrafish exhibit increased locomotor sensitivity to PTZ and elevated expression of epileptogenesis-related genes C_LIO_LIkcnb1-/- larvae show spontaneous and provoked epileptiform-like electrographic activity associated with disrupted GABA regulation C_LI ObjectiveKCNB1 encodes an -subunit of the delayed-rectifier voltage-dependent potassium channel Kv2.1. De novo pathogenic variants of KCNB1 have been linked to developmental and epileptic encephalopathies (DEE), diagnosed in early childhood and sharing limited treatment options. Loss-of-function (LOF) of KCNB1 with dominant negative effects has been proposed as the pathogenic mechanism in these disorders. Here, we aim to characterize a knock-out (KO) zebrafish line targeting kcnb1 (kcnb1+/- and kcnb1-/-) for investigating DEEs. MethodsThis study presents the phenotypic analysis of a kcnb1 knock-out zebrafish model, obtained by CRISPR/Cas9 mutagenesis. Through a combination of immunohistochemistry, behavioral assays, electrophysiological recordings, and neurotransmitter quantifications, we have characterized the expression, function, and impact of this kcnb1 LOF model at early stages of development. ResultsIn wild-type larval zebrafish, kcnb1 was found in various regions of the central nervous system and in diverse cell subtypes including neurons, oligodendrocytes and microglial cells. Both kcnb1+/- and kcnb1-/- zebrafish displayed impaired swimming behavior and "epilepsy-like" features that persisted through embryonic and larval development, with variable severity. When exposed to the chemoconvulsant pentylenetetrazol (PTZ), both mutant models showed elevated locomotor activity. In addition, PTZ-exposed kcnb1-/- larvae exhibited higher bdnf mRNA expression and activated c-Fos positive neurons in the telencephalon. This same model presents spontaneous and provoked epileptiform-like electrographic activity associated with disrupted GABA regulation. In this KO model, neuronal circuit organization remained unaffected. SignificanceWe conclude that kcnb1 knock-out in zebrafish leads to early-onset phenotypic features reminiscent of DEEs, affecting neuronal functions and primarily inhibitory pathways in developing embryonic and larval brains. This study highlights the relevance of this model for investigating developmental neuronal signaling pathways in KCNB1-related DEEs.

neuroscience↗

bioRxiv2023

Abnormal autophagy is a critical mechanism in TANGO2-related rhabdomyolysis

Patients with pathogenic variants in the TANGO2 gene suffer from severe and recurrent rhabdomyolysis (RM) episodes precipitated by fasting. Since starvation promotes autophagy induction, we wondered whether TANGO2-related muscle symptoms result from autophagy insufficiency to meet cellular demands in stress conditions. Autophagy functioning was analyzed in vitro, in primary skeletal muscle cells from TANGO2 patients in basal and fasting conditions. In addition, we developed a tango2 morphant zebrafish model to assess the effect of tango2 knockdown (KD) on locomotor function and autophagy efficiency in vivo. We report that TANGO2 mutations are associated with decreased LC3-II levels upon starvation in primary muscle cells, but not in fibroblasts. In zebrafish larvae, tango2 knockdown induces locomotor defects characterized by reduced evoked movements which are exacerbated by exposure to atorvastatin, a compound known to cause RM. Importantly, RM features of tango2 KD are also associated with autophagy and mitophagy defects in zebrafish. Calpeptin treatment, a known activator of autophagy, is sufficient to rescue the locomotor properties, thanks to its beneficial effect on autophagy functioning in zebrafish and independently to its effect on calpain activity. LC3-II levels of primary muscle cells of TANGO2 patients are also improved by calpeptin treatment. Overall, we demonstrate that TANGO2 plays an important role in autophagy, and that autophagy efficiency is critical to prevent RM, thus giving rise to new therapeutic perspectives in the prevention of these life-threatening episodes in TANGO2 pathology.

cell biology↗

bioRxiv2021

Poly(A)-binding protein is an ataxin-2 chaperone that emulsifies biomolecular condensates.

Biomolecular condensation underlies the biogenesis of an expanding array of membraneless assemblies, including stress granules (SGs) which form under a variety of cellular stresses. Advances have been made in understanding the molecular grammar that dictates the behavior of a few key scaffold proteins that make up these phases but how the partitioning of hundreds of other SG proteins is regulated remains largely unresolved. While investigating the rules that govern the condensation of ataxin-2, a SG protein implicated in neurodegenerative disease, we unexpectedly identified a short 14aa sequence that acts as an ataxin-2 condensation switch and is conserved across the eukaryote lineage. We identify poly(A)-binding proteins as unconventional RNA-dependent chaperones that control this regulatory switch. Our results uncover a hierarchy of cis and trans interactions that fine-tune ataxin-2 condensation and reveal a new molecular function for ancient poly(A)-binding proteins as emulsifiers of biomolecular condensate proteins. These findings may inspire novel approaches to therapeutically target aberrant phases in disease.

cell biology↗

bioRxiv2020

Functional characterization of Neurofilament Light b splicing and misbalance in zebrafish

Neurofilaments (NFs), a major cytoskeletal component of motor neurons, play a key role in their differentiation, establishment and maintenance of their morphology and mechanical strength. The de novo assembly of these neuronal intermediate filaments requires the presence of the neurofilament light subunit, NEFL, which expression is reduced in motor neurons in Amyotrophic Lateral Sclerosis (ALS). This study used zebrafish as a model to characterize the NEFL homologue neflb, which encodes two different isoforms via splicing of the primary transcript (neflbE4 and neflbE3). In vivo imaging showed that neflb is crucial for proper neuronal development, and that disrupting the balance between its two isoforms specifically affects NF assembly and motor axon growth, with resulting motor deficits. This equilibrium is also disrupted upon partial depletion of TDP-43, a RNA binding protein that is mislocalized into cytoplasmic inclusions in ALS. The study supports interaction of NEFL expression and splicing with TDP-43 in a common pathway, both biologically and pathogenetically.

neuroscience↗