Biology subjects

Kalcheim, C.

Publications and source records attributed to Kalcheim, C..

5 recordsLinked to original sources

bioRxiv2025

Dynamic Functional Pathway Development in Type 1 Spinal Interneurons: Stage-specific roles of retinoic acid activity

BackgrounddI1 spinal interneuron development requires precise coordination of proliferation, differentiation, and migration, yet the molecular and cellular properties underlying the temporal coupling of these processes remain unresolved. ResultsSingle-cell RNA sequencing of the E4 quail neural tube under control and retinoic acid (RA)-deprived conditions generated a high-resolution atlas spanning a continuous trajectory from dorsal progenitors to differentiated neurons. Pseudotime and RNA velocity analyses revealed transitions consistent with medial-to-lateral displacement in vivo. Early progenitors expressed cell cycle and BMP-associated programs, followed by induction of cytoskeletal and extracellular matrix remodelling genes and later activation of pan-neuronal and synaptic pathways. BMP signaling declined during differentiation through upregulation of BMP antagonists, whereas apoptosis-related genes were poised during specification without overt cell death. Autophagy-associated genes peaked during differentiation. Disruption of RA signaling by dominant-negative RAR403 uncovered stage-specific functions of RA. In progenitors, RA inhibition increased proliferation. In differentiating neurons, it upregulated BARHL1/2 and enhanced ion channel, synaptic, cytoskeletal, and extracellular matrix gene programs. RA inhibition also impaired ventral displacement of BarHL2-positive dI1 neurons, causing abnormal positioning. ConclusionsThese findings define a dI1-centered single-cell atlas of the progenitor-to-neuron transition and identify stage-specific roles for RA in dorsal hinterneuron maturation. Key findingsO_LIWe provide a high-resolution single-cell atlas of dI1 spinal interneuron development in the E4 quail neural tube, analysis of this single stage reveals a continuous trajectory from proliferative dorsal progenitors to differentiated neurons. C_LIO_LIDevelopment along this trajectory is tightly staged: early cells are dominated by cell-cycle and progenitor-maintenance programs, while later cells progressively activate cytoskeletal, extracellular matrix, axon-guidance, synaptic, and mature neuronal pathways. C_LIO_LIBMP signaling is strongest in early dI1 progenitors and is progressively attenuated during differentiation through the induction of BMP inhibitors, supporting a stage-specific role for BMP in early specification rather than late maturation. C_LIO_LIRetinoic acid acts in a stage-dependent manner: RA inhibition increases progenitor proliferation, alters differentiation-associated gene programs, and upregulates neuronal maturation, ion-channel, and synaptic genes in differentiating neurons. C_LIO_LIRepressing RA signaling causes BarHL2-positive dI1 neurons to fail their normal ventral migration. Instead, they accumulate near their dorsal origin and show ectopic localization, indicating that RA is required for proper migration. C_LI

developmental biology↗

bioRxiv2024

Retinoic acid, an essential component of the RP organizer, promotes the spatio-temporal segregation of dorsal neural fates

Dorsal neural tube-derived retinoic acid promotes the end of neural crest production and transition into a definitive roof plate. Here we analyze how this impacts the segregation of central and peripheral lineages, a process essential for tissue patterning and function. Localized in-ovo inhibition of retinoic acid activity followed by single cell transcriptomics unraveled a comprehensive list of differentially expressed genes relevant to these processes. Importantly, progenitors co-expressed neural crest, roof plate and dI1 interneuron markers indicating a failure in proper lineage segregation. Furthermore, we found that separation between roof plate and dI1 interneurons is mediated by Notch activity downstream of retinoic acid, highlighting their critical role in establishing the roof plate-dI1 boundary. Within the peripheral branch, where absence of retinoic acid resulted in neural crest production and emigration extending into the roof plate stage, sensory progenitors failed to separate from melanocytes leading to formation of a common glia-melanocyte cell with aberrant migratory patterns. Together, we uncover and characterize a molecular mechanism responsible for segregation of dorsal neural fates during development.

developmental biology↗

bioRxiv2021

Completion of neural crest cell production and emigration is regulated by retinoic acid-dependent inhibition of BMP signaling

Production and emigration of neural crest cells is a transient process followed by the emergence of the definitive roof plate. The mechanisms regulating the end of neural crest ontogeny are poorly understood. Whereas early crest development is stimulated by mesoderm-derived retinoic acid, we report that the end of the neural crest period is regulated by retinoic acid synthesized in the dorsal neural tube. Inhibition of retinoic acid signaling in the neural tube prevents the normal upregulation of BMP inhibitors in the nascent roof plate and prolongs the period of BMP responsiveness which otherwise ceases close to roof plate establishment. Consequently, neural crest production and emigration are extended well into the roof plate stage. In turn, extending the activity of neural crest-specific genes inhibits the onset of retinoic acid synthesis in roof plate suggesting a mutual repressive interaction between neural crest and roof plate traits. Although several roof plate-specific genes are normally expressed in the absence of retinoic acid signaling, roof plate and crest markers are co-expressed in single cells and this domain also contains dorsal interneurons. Hence, the cellular and molecular architecture of the roof plate is compromised. Collectively, our results demonstrate that neural tube-derived retinoic acid, via inhibition of BMP signaling, is an essential factor responsible for the end of neural crest generation and the proper segregation of dorsal neural lineages.

developmental biology↗

bioRxiv2020

Notch signaling is a critical initiator of roof plate formation as revealed by the use of RNA profiling of the dorsal neural tube

The factors underlying establishment of the definitive roof plate (RP) and its segregation from neural crest (NC) and interneurons are unknown. We performed transcriptome analysis at trunk levels of quail embryos comparing the dorsal neural tube at premigratory NC and RP stages. This unraveled molecular heterogeneity between NC and RP stages, and within the RP itself. By implementing these genes, we asked whether Notch signaling is involved in RP development. First, we observed that Notch is active at the RP-interneuron interface. Furthermore, gain and loss of Notch function in quail and mouse embryos, respectively, revealed no effect on early NC behavior. Constitutive Notch activation caused a local downregulation of RP markers with a concomitant development of dI1 interneurons, as well as an ectopic upregulation of RP markers in the interneuron domain. Reciprocally, in mice lacking Notch activity both the RP and dI1 interneurons failed to form and this was associated with expansion of the dI2 population. Collectively, our results offer a new resource for defining specific cell types, and provide evidence that Notch is required to establish the definitive RP, and to determine the choice between RP and interneuron fates, but not the segregation of RP from NC. Summary statementA new set of genes involved in Notch-dependent roof plate formation is unraveled by transcriptome analysis.

developmental biology↗

bioRxiv2019

Neural tube development depends on notochord-derived Sonic hedgehog released into the sclerotome

Sonic hedgehog (Shh), produced in notochord and floor plate, is necessary both for neural and mesodermal development. To reach the myotome, Shh has to traverse the sclerotome. By loss and gain of Shh function, and floor plate deletions, we report that sclerotomal Shh is also necessary for neural tube development. Reducing the amount of Shh in sclerotome by membrane-tethered hedgehog-interacting protein or by Patched1, but not by dominant active Patched, decreased motoneuron numbers while also compromising myotome differentiation. These effects were a specific and direct consequence of reducing Shh. In addition, grafting notochords in a basal, but not apical location vis-a-vis the tube, profoundly affected motoneuron development, suggesting that initial ligand presentation occurs at the basal side of epithelia corresponding to the sclerotome-neural tube interface.\n\nCollectively, our results reveal that the sclerotome is a potential site of a Shh gradient that coordinates development of mesodermal and neural progenitors.\n\nSummary statementShh that transits through the sclerotome is presented to the neuroepithelium from its basal aspect to affect motoneuron development.

developmental biology↗