Biology subjects

Gillotay, P.

Publications and source records attributed to Gillotay, P..

8 recordsLinked to original sources

bioRxiv2026

Microenvironmental and transcriptional determinants of the robustness of spinal cord regeneration in zebrafish

Unlike mammals, adult zebrafish can regenerate spinal cord tissue after complete transection. We report here that a substantial proportion of zebrafish receiving this injury fail to regain locomotor function, and we analyzed this critically. After stratifying large cohorts of injured animals according to functional recovery, we directly compared recovered and non-recovered individuals at matched post-injury stages. We find that permanently paralyzed animals form a tissue bridge across the lesion yet exhibit impaired axonal invasion and regrowth to downstream targets. Integrative single-nucleus profiling, spatial transcriptomics, and regulatory network analyses revealed that regenerative success is characterized by the coordinated establishment of a permissive multicellular regenerative state rather than major differences in cellular composition. Furthermore, we find that regenerative failure involves persistent non-permissive regulatory programs, including extracellular matrix remodeling, impaired adaptive T-cell activation, and reduced axon growth-associated signaling. Gene regulatory network and cellular trajectory analyses identified candidate transcriptional regulators that could not be resolved through differential expression analysis alone. Functional CRISPR F0 screening of several of these candidates identified hoxb5a as a critical determinant of regenerative outcome. Stable hoxb5a mutants exhibited impaired swimming recovery, disrupted bridge formation, and limited axon bridging after spinal cord injury. Together, our findings reveal key differences between success and failure in a model of elevated regenerative capacity, with hoxb5a acting as a regulator linking multicellular tissue remodeling to axonal regeneration and functional recovery.

developmental biology↗

bioRxiv2022

Foxe1 orchestrates thyroid and lung cell lineage divergence in mouse stem cell-derived organoids

Patterning of endoderm into lung and thyroid lineages depends upon a correct early expression of a homeobox domain-containing transcription factor, Nkx2-1. However, the gene networks distinguishing the differentiation of those lineages remain largely unknown. In the present work, by using mouse embryonic stem cell lines, single-cell RNA sequencing, and transcriptomic and chromatin accessibility profiling, we show that knockout of Foxe1 drastically impairs Nkx2-1+ cells differentiation and maturation into thyroid follicular-like cells. Concomitantly, a subset of Foxe1 null/Nkx2-1+ cells have a remarkable ability in vitro to undergo a lung epithelial differentiation program and form lung-like organoids harboring cells transcriptionally similar with mouse fetal airway and alveolar cell types. These results demonstrate, for the first time, lung lineage derivation at the expense of thyroid lineage, by a simple removal of a transcription factor, and provide insights into the intricated mechanisms of fate decisions of endodermal cell types. Highlights- Forward programming of mESCs with transient Nkx2-1 and Pax8 overexpression, followed by c-AMP treatment, leads to differentiation of functional thyroid follicles in vitro; - In absence of Foxe1, thyroid follicle-like structures, derived from mESCs, are scarce and non-functional; - Concomitantly, a subset of Nkx2-1-expressing cells generated from Foxe1KO mESCs spontaneously form lung organoids containing multiple differentiated lung cell types; - ATACseq analyses show higher chromatin remodeling in Nkx2-1-expressing cells in control compared to Foxe1KO cells, especially for genes involved in thyroid maturation and maintenance of the 3D structure of the follicle.

developmental biology↗

bioRxiv2022

Nrf2 promotes thyroid development and hormone synthesis.

In all vertebrates, the function of the thyroid gland is to capture iodide to synthesize hormones that act on almost all tissues and are essential for normal growth and metabolism. Thyroid hormone production is a multistep process that ultimately leads to the coupling of iodine to thyroglobulin, the matrix glycoprotein of hormone synthesis. This coupling is dependent on H2O2, the accumulation of which in thyroid tissue, a true iodine trap, causes a basal level of oxidative stress higher than in other tissues, which, if left unchecked, leads to cellular damage. The thyroid has efficient antioxidant and detoxifying enzymes that help it resist H202-induced oxidative stress and maintain the homeostasis necessary for hormone synthesis. By regulating the expression of genes involved in cellular detoxification processes, the transcription factor NRF2 acts as a master regulator of the cellular defense system against oxidative stress. Using zebrafish embryos and mouse thyroid organoids, we show here that direct control of thyroglobulin expression by NRF2 is an evolutionarily conserved chore mechanism in vertebrates to complete the necessary molecular defense arsenal against oxidative stress in thyroid tissue.

developmental biology↗

bioRxiv2021

Transplantable human thyroid organoids generated from embryonic stem cells to rescue hypothyroidism

The thyroid gland captures iodide in order to synthesize hormones that act on almost all tissues and are essential for normal growth and metabolism. Low plasma levels of thyroid hormones lead to hypothyroidism, which is one of the most common disorder in humans and is not always satisfactorily treated by lifelong hormone replacement. Therefore, in addition to the lack of in vitro tractable models to study human thyroid development, differentiation and maturation, functional human thyroid organoids could pave the way to explore new therapeutic approaches. Here we report the first transplantable thyroid organoids derived from human embryonic stem cells capable of restoring plasma thyroid hormone to athyreotic mice as a proof of concept for future therapeutic development.

developmental biology↗

bioRxiv2020

Timed mesodermal FGF and BMP govern the multi-step thyroid specification

Thyroid tissue is the site for de novo synthesis of thyroid hormones which are essential for vertebrate development and growth. Defects in embryonic thyroid morphogenesis are a predominant cause for congenital thyroid diseases but the molecular pathomechanisms are incompletely understood. The first molecularly recognizable step of thyroid development is the specification of thyroid precursors at a defined position in the anterior foregut endoderm. While recent studies identified FGF and BMP pathways as critical signaling factors for thyroid specification, the interplay between extrinsic signaling cues and thyroid transcription factor expression remained elusive. Here, we used zebrafish embryos to decipher the dynamics of thyroid transcription factor induction in relation to FGF and BMP signaling activities in pharyngeal endoderm. We first identified a previously unrecognized endodermal thyroid progenitor cell population expressing Pax2a but not Nkx2.4b. This cell population is characterized by enhanced FGF signaling but initially lacks detectable BMP signaling. A subpopulation of Pax2a-expressing progenitors differentiates subsequently into thyroid lineage-committed precursor cells co-expressing Pax2a and Nkx2.4b. We next combined pharmacological approaches with genetic models permitting inhibition or ectopic overactivation of signaling pathways to timely manipulate FGF and BMP activities. These experiments support a model where FGF signaling primarily regulates Pax2a expression whereas BMP signaling has dual functions in regulation of both Pax2a and Nkx2.4b expression. Collectively, our data allow us to formulate a refined model of thyroid cell specification from foregut endoderm.

developmental biology↗

bioRxiv2020

Single-cell transcriptome analysis reveals cell-cell communication and thyrocyte diversity in the zebrafish thyroid gland

The thyroid gland regulates growth and metabolism via production of thyroid hormone in follicles composed of thyrocytes. So far, thyrocytes have been assumed to be a homogenous population. To uncover genetic heterogeneity in the thyrocyte population, and molecularly characterize the non-thyrocyte cells surrounding the follicle, we developed a single-cell transcriptome atlas of the zebrafish thyroid gland. The 6249-cell atlas includes profiles of thyrocytes, blood vessels, lymphatic vessels, immune cells and fibroblasts. Further, the thyrocytes could be split into two sub-populations with unique transcriptional signature, including differential expression of the transcription factor pax2a. To validate thyrocyte heterogeneity, we generated a CRISPR/Cas9-based pax2a knock-in line, which demonstrated specific pax2a expression in the thyrocytes. However, a population of pax2a-low mature thyrocytes interspersed within individual follicles could be distinguished, corroborating heterogeneity within the thyrocyte population. Our results identify and validate transcriptional differences within the nominally homogenous thyrocyte population. One-line summarySingle-cell analysis uncovers latent heterogeneity in thyroid follicular cells. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/891630v2_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@f6df3org.highwire.dtl.DTLVardef@fb8c59org.highwire.dtl.DTLVardef@bb5be1org.highwire.dtl.DTLVardef@8d174e_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2019

Enhanced canonical Wnt signaling during early zebrafish development perturbs the interaction of cardiac mesoderm and pharyngeal endoderm and causes thyroid specification defects

BackgroundCongenital hypothyroidism (CH) due to thyroid dysgenesis is a frequent congenital endocrine disorder for which the molecular mechanisms remain unresolved in the far majority of cases. This situation reflects in part our still limited knowledge about the mechanisms involved in the early steps of thyroid specification from the endoderm, in particular the extrinsic signaling cues that regulate foregut endoderm patterning. In this study, we used small molecules and genetic zebrafish models to characterize the role of various signaling pathways in thyroid specification. MethodsWe treated zebrafish embryos during different developmental periods with small molecule compounds known to modulate the activity of Wnt signaling pathway and observed effects in thyroid, endoderm and cardiovascular development using whole mount in situ hybridization and transgenic fluorescent reporter models. We used an antisense morpholino to create a zebrafish acardiac model. For thyroid rescue experiments, BMP pathway induction in zebrafish embryos was obtained by using heatshock inducible transgenic lines. ResultsInterestingly, combined analyses of thyroid and cardiovascular development revealed that overactivation of Wnt signaling during early development leads to impaired thyroid specification concurrent with severe defects in the cardiac specification. When using a model of morpholino-induced blockage of cardiomyocyte differentiation, a similar correlation was observed, suggesting that defective signaling between cardiac mesoderm and endodermal thyroid precursors contributes to thyroid specification impairment. Rescue experiments through transient overactivation of BMP signaling could partially restore thyroid specification in models with defective cardiac development. ConclusionCollectively, our results indicate that BMP signaling is critically required for thyroid cell specification and identify cardiac mesoderm as a likely source of BMP signals.

developmental biology↗

bioRxiv2019

Small Molecule Screening in Zebrafish Embryos Identifies Signaling Pathways Regulating Early Thyroid Development

BackgroundDefects in embryonic development of the thyroid gland are a major cause for congenital hypothyroidism in human newborns but the underlying molecular mechanisms are still poorly understood. Organ development relies on a tightly regulated interplay between extrinsic signaling cues and cell intrinsic factors. At present, however, there is limited knowledge about the specific extrinsic signaling cues that regulate foregut endoderm patterning, thyroid cell specification and subsequent morphogenetic processes in thyroid development. MethodsTo begin to address this problem in a systematic way, we used zebrafish embryos to perform a series of in vivo phenotype-driven chemical genetic screens to identify signaling cues regulating early thyroid development. For this purpose, we treated zebrafish embryos during different developmental periods with a panel of small molecule compounds known to manipulate the activity of major signaling pathways and scored phenotypic deviations in thyroid, endoderm and cardiovascular development using whole mount in situ hybridization and transgenic fluorescent reporter models. ResultsSystematic assessment of drugged embryos recovered a range of thyroid phenotypes including expansion, reduction or lack of the early thyroid anlage, defective thyroid budding as well as hypoplastic, enlarged or overtly disorganized presentation of the thyroid primordium after budding. Our pharmacological screening identified BMP and FGF signaling as key factors for thyroid specification and early thyroid organogenesis, highlight the importance of low Wnt activities during early development for thyroid specification and implicate drug-induced cardiac and vascular anomalies as likely indirect mechanisms causing various forms of thyroid dysgenesis. ConclusionsBy integrating the outcome of our screening efforts with previously available information from other model organisms including Xenopus, chicken and mouse, we conclude that signaling cues regulating thyroid development appear broadly conserved across vertebrates. We therefore expect that observations made in zebrafish can inform mammalian models of thyroid organogenesis to further our understanding of the molecular mechanisms of congenital thyroid diseases.

developmental biology↗