Biology subjects

Eschstruth, A.

Publications and source records attributed to Eschstruth, A..

7 recordsLinked to original sources

bioRxiv2026

Spinal lumen remodeling under the control of Gli signaling mechanically drives roof plate cells extension

Morphogenesis often requires different cell types to coordinate their behaviors for an harmonious developpement. How these different cell type behaviors are synchronized within and between tissues remains one of the important questions to fully understand morphogenesis. We used the zebrafish developing spinal cord to study this question. At later stages of neurogenesis, the lumen of the neural tube remodels, by reducing its height dramatically to form the persisting ventral central canal, a morphogenetic process conserved in vertebrates. By combining genetics, cell signaling manipulation with antagonist drugs and high-resolution in vivo live imaging, we better characterised the dynamics and control of this remodeling process. We showed that the lumen retraction depends on Gli activity regulation, a downstream effector of the Shh morphogen signal. We further established that the lumen retraction is instrumental in the cellular elongation of spinal roof plate cells, a population that forms the ceiling of the spinal cord lumen. Our work therefore establishes that the Gli transcriptional regulators under the control of long-range morphogen Shh control lumen retraction and that this retraction is a key driver of the roof plate cells extension.

developmental biology↗

bioRxiv2025

Dynamic interactions between epithelial skin cells and a sensory cavity sculpt the growing olfactory orifice

During morphogenesis and in pathological conditions, gaps can form in the plane of epithelial barriers upon cellular forces that disrupt intercellular junctions. How the size of these epithelial holes further increases over time and what sets their shape remain poorly understood. Here we analyze the formation of the olfactory orifice (the nostril) in zebrafish, which opens and grows in the skin epithelium above a rosette of olfactory placode cells, allowing the sensory neurons to directly access odor cues. Using quantitative imaging and tissue-specific perturbations, we analyzed the dynamic remodeling of skin cells allowing the expansion of the orifice edge. We identified the sensory cavity located in the center of the placodal rosette as a crucial player that sets the size of the growing epithelial hole in the skin. We further showed that fine-tuning of actomyosin contractility within each tissue (skin and sensory cavity) exerts non-autonomous effects on the neighboring tissue, thereby shaping the nostril structure. This study uncovers dynamic cell behaviors and reciprocal tissue-tissue interplay that control the growth and shape of an epithelial hole in vivo.

developmental biology↗

bioRxiv2024

A DIFFERENTIAL REQUIREMENT FOR THE CILIOPATHY GENE RPGRIP1L IN HUMAN AND MOUSE SPINAL PROGENITOR FATE SPECIFICATION

Studying developmental processes in the context of the human central nervous system is essential to understand neurodevelopmental diseases. In this paper we perform a comparative functional study of the ciliopathy gene RPGRIP1L in human and mouse spinal development using in vitro 3D differentiation of pluripotent stem cells. RPGRIP1L, a causal gene of severe neurodevelopmental ciliopathies such as Joubert and Meckel syndromes, encodes a scaffolding protein of the ciliary transition zone involved in ciliary gating. Previous work has identified a major role for Rpgrip1l in mouse brain and spinal cord development, via controlling the Sonic Hedgehog (SHH)/GLI pathway. We show that spinal organoids derived from Rpgrip1l mutant mouse embryonic stem cells faithfully recapitulate the loss of motoneurons and the strong reduction of SHH signaling observed in the mutant mice. In contrast, human induced pluripotent stem cells mutant for RPGRIP1L produce motoneurons and activate the SHH pathway at levels similar to wild types, a property shared by human iPSCs mutant for another ciliopathy gene TMEM67. Moreover, we show that, in human RPGRIP1L mutant organoids, motoneurons acquire a more anterior identity, expressing HOX genes and other proteins normally present in the hindbrain and cervical spinal cord while motoneurons from wild type organoids strictly display a caudal brachial identity. By performing a temporal transcriptome analysis throughout the differentiation process, we find that the antero-posterior specification defect arises in early axial progenitors and correlates with the loss of cilia in these cells. Thus, this study uncovers distinct functions in humans and mice for ciliopathy proteins and a novel role for RPGRIP1L in human spinal antero-posterior patterning. These findings have important implications for understanding the role of cilia in human spinal cord development and the pathogenic mechanisms of neurodevelopmental ciliopathies.

developmental biology↗

bioRxiv2024

ASTROGLIOSIS AND NEUROINFLAMMATION UNDERLIE SCOLIOSIS UPON CILIA DYSFUNCTION

Cilia defects lead to scoliosis in zebrafish, but the underlying pathogenic mechanisms are poorly understood and may diverge depending on the mutated gene. Here, we dissected the mechanisms of scoliosis onset in a zebrafish mutant for the rpgrip1l gene encoding a ciliary transition zone protein. rpgrip1l mutant fish developed scoliosis with near-total penetrance but asynchronous onset in juveniles. Taking advantage of this asynchrony, we found that curvature onset was preceded by ventricle dilations and was concomitant to the perturbation of Reissner fiber polymerization and to the loss of multiciliated tufts around the subcommissural organ. Rescue experiments showed that Rpgrip1l was exclusively required in foxj1a-expressing cells to prevent axis curvature. Genetic interactions investigations ruled out Urp1/2 levels as a main driver of scoliosis in rpgrip1 mutants. Transcriptomic and proteomic studies identified neuroinflammation associated with increased Annexin levels as a potential mechanism of scoliosis development in rpgrip1l juveniles. Investigating the cell types associated with annexin2 over-expression, we uncovered astrogliosis, arising in glial cells surrounding the diencephalic and rhombencephalic ventricles just before scoliosis onset and increasing with time in severity. Anti-inflammatory drug treatment reduced scoliosis penetrance and severity and this correlated with reduced astrogliosis and macrophage/microglia enrichment around the diencephalic ventricle. Mutation of the cep290 gene encoding another transition zone protein also associated astrogliosis with scoliosis. Thus, we propose astrogliosis induced by perturbed ventricular homeostasis and associated with immune cell activation as a novel pathogenic mechanism of zebrafish scoliosis caused by cilia dysfunction.

developmental biology↗

bioRxiv2023

Cerebellar granular neuron progenitors exit their germinative niche via Barhl1 mediated silencing of T-Cell Factor transcriptional activity

SUMMARYT-Cell Factors (TCFs) are the main transcriptional effectors of Wnt/{beta}-catenin signaling. TCF responsiveness is a hallmark of self-renewal in mouse embryonic, and adult, neural stem cells (NSC). However, in vivo contribution(s) of TCF activities in long-lived NSCs are poorly understood. Granule neuron progenitors (GNP) in the upper rhombic lip (URL) are long-lived NSCs which express Atoh1 and generate cerebellar granule neurons. Using functional and transcriptomic approaches in amphibian, we demonstrate that TCFs are active in the URL, and are strictly necessary for the emergence and maintenance of the GNP germinative zone. We identify BarH-like 1 (Barhl1), a direct target of Atoh1, as a gate keeper for GNP exit from the URL, through silencing of TCF transcriptional activity. Our transcriptomic and in silico analysis identifies Barhl1/TCF URL target genes, and confirms our functional data. Our study provides in vivo evidence that inhibition of TCF repressive activity is necessary for maintenance of the URL, a long-lived neural germinative niche.

developmental biology↗

bioRxiv2022

Retinoic acid control of pax8 during renal specification of Xenopus pronephros involves hox and meis3

Development of the Xenopus pronephros relies on renal precursors grouped at neurula stage into a specific region of dorso-lateral mesoderm called the kidney field. Formation of the kidney field at early neurula stage is dependent on retinoic (RA) signaling acting upstream of renal master transcriptional regulators such as pax8 or lhx1. Although lhx1 might be a direct target of RA-mediated transcriptional activation in the kidney field, how RA controls the emergence of the kidney field remains poorly understood. In order to better understand RA control of renal specification of the kidney field, we have performed a transcriptomic profiling of genes affected by RA disruption in lateral mesoderm explants isolated prior to the emergence of the kidney field and cultured at different time points until early neurula stage. Besides genes directly involved in pronephric development (pax8, lhx1, osr2, mecom), hox (hoxa1, a3, b3, b4, c5 and d1) and the hox co-factor meis3 appear as a prominent group of genes encoding transcription factors (TFs) downstream of RA. Supporting the idea of a role of meis3 in the kidney field, we have observed that meis3 depletion results in a severe inhibition of pax8 expression in the kidney field. Meis3 depletion only marginally affects expression of lhx1 and aldh1a2 suggesting that meis3 principally acts upstream of pax8. Further arguing for a role of meis3 and hox in the control of pax8, expression of a combination of meis3, hoxb4 and pbx1 in animal caps induces pax8 expression, but not that of lhx1. The same combination of TFs is also able to transactivate a previously identified pax8 enhancer, Pax8-CNS1. Mutagenesis of potential PBX-Hox binding motifs present in Pax8-CNS1 further allows to identify two of them that are necessary for transactivation. Finally, we have tested deletions of regulatory sequences in reporter assays with a previously characterized transgene encompassing 36.5 kb of the X. tropicalis pax8 gene that allows expression of a truncated pax8-GFP fusion protein recapitulating endogenous pax8 expression. This transgene includes three conserved pax8 enhancers, Pax8-CNS1, Pax8-CNS2 and Pax8-CNS3. Deletion of Pax8-CNS1 alone does not affect reporter expression, but deletion of a 3.5kb region encompassing Pax8-CNS1 and Pax8-CNS2 results in a severe inhibition of reporter expression both in the otic placode and kidney field domains.

developmental biology↗

bioRxiv2022

Hnf1b renal expression directed by a distal enhancer responsive to Pax8

Xenopus provides a simple and efficient model system to study nephrogenesis and explore the mechanisms causing renal developmental defects in human. Hnf1b (hepatocyte nuclear factor 1 homeobox b), a gene whose mutations are the most commonly identified genetic cause of developmental kidney disease, is required for the acquisition of a proximo-intermediate nephron segment in Xenopus as well as in mouse. Genetic networks involved in Hnf1b expression during kidney development remain poorly understood. We decided to explore the transcriptional regulation of Hnf1b in the developing Xenopus pronephros and mammalian renal cells. Using phylogenetic footprinting, we identified an evolutionary conserved sequence (CNS1) located several kilobases (kb) upstream the Hnf1b transcription start and harboring epigenomic marks characteristics of a distal enhancer in embryonic and adult renal cells in mammals. By means of functional expression assays in Xenopus and mammalian renal cell lines we showed that CNS1 displays enhancer activity in renal tissue. Using CRISPR/cas9 editing in Xenopus tropicalis, we demonstrated the in vivo functional relevance of CNS1 in driving hnf1b expression in the pronephros. We further showed the importance of Pax8-CNS1 interaction for CNS1 enhancer activity allowing us to conclude that Hnf1b is a direct target of Pax8. Our work identified for the first time a Hnf1b renal specific enhancer and may open important perspectives into the diagnosis for congenital kidney anomalies in human, as well as modeling HNF1B-related diseases.

developmental biology↗