Biology subjects

Schilling, T. F.

Publications and source records attributed to Schilling, T. F..

6 recordsLinked to original sources

bioRxiv2025

The ratio of Wnt signaling activity to Sox2 transcription factor levels predicts neuromesodermal fate potential

Neuromesodermal progenitors (NMPs) are a vertebrate cell type that contribute descendants to both the spinal cord and the mesoderm. The undifferentiated bipotential NMP state is maintained when both Wnt signaling is active and Sox2 is present. We used transgenic reporter lines to live-image both Wnt activity and Sox2 levels in NMPs and observed a unique cellular ratio in NMPs compared to NMP-derived mesoderm or neural tissue. We used this unique signature to identify the previously unknown anatomical position of a progenitor population that gives rise to the midline tissues of the floor plate of the spinal cord and the mesodermal notochord. Thus, quantification of the active Wnt signaling to Sox2 ratio can be used to predict and identify cells with neuromesodermal potential. We also developed the auxin inducible degron 2 system for use in zebrafish to test the temporal role that Sox2 plays during midline formation. We found ectopic Sox2 in the presence of Wnt activity holds cells in the undifferentiated floor plate/notochord progenitor state, and that degradation of the ectopic Sox2 is required for cells to adopt a notochord fate.

developmental biology↗

bioRxiv2023

Cohesin composition and dosage independently affect early development in zebrafish

Cohesin, a chromatin-associated protein complex with four core subunits (Smc1a, Smc3, Rad21 and either Stag1 or 2), has a central role in cell proliferation and gene expression in metazoans. Human developmental disorders termed "cohesinopathies" are characterised by germline mutations in cohesin or its regulators that do not entirely eliminate cohesin function. However, it is not clear if mutations in individual cohesin subunits have independent developmental consequences. Here we show that zebrafish rad21 or stag2b mutants independently influence embryonic tailbud development. Both mutants have altered mesoderm induction, but only homozygous or heterozygous rad21 mutation affects cell cycle gene expression. stag2b mutants have narrower notochords and reduced Wnt signaling in neuromesodermal progenitors as revealed by single cell RNA-sequencing. Stimulation of Wnt signaling rescues transcription and morphology in stag2b, but not rad21 mutants. Our results suggest that mutations altering the quantity versus composition of cohesin have independent developmental consequences, with implications for the understanding and management of cohesinopathies. Summary StatementViable zebrafish mutants show that cohesin complex quantity versus composition lead to different transcriptional and developmental outcomes in the early embryo.

developmental biology↗

bioRxiv2022

Transcriptome profiling of tendon fibroblasts at the onset of embryonic muscle contraction reveals novel force-responsive genes

Mechanical forces play a critical role in tendon development and function, influencing cell behavior through mechanotransduction signaling pathways and subsequent extracellular matrix (ECM) remodeling. Here we investigate the molecular mechanisms by which tenocytes in developing zebrafish embryos respond to muscle contraction forces during the onset of swimming and cranial muscle activity. Using genome-wide bulk RNA sequencing of FAC-sorted tenocytes we identify novel tenocyte markers and genes involved in tendon mechanotransduction. Embryonic tendons show dramatic changes in expression of Matrix Remodeling Associated 5b (mxra5b), Matrilin 1 (matn1), and the transcription factor Kruppel-like factor 2a (klf2a), as muscles start to contract. Using embryos paralyzed either by loss of muscle contractility or neuromuscular stimulation we confirm that muscle contractile forces influence the spatial and temporal expression patterns of all three genes. Quantification of these gene expression changes across tenocytes at multiple tendon entheses and myotendinous junctions reveals that their responses depend on force intensity, duration and tissue stiffness. These force-dependent feedback mechanisms in tendons, particularly in the ECM, have important implications for improved treatments of tendon injuries and atrophy.

developmental biology↗

bioRxiv2021

Multiple morphogens and rapid elongation promote segmental patterning during development

The vertebrate hindbrain is segmented into rhombomeres (r) initially defined by distinct domains of gene expression. Previous studies have shown that noise-induced gene regulation and cell sorting are critical for the sharpening of rhombomere boundaries, which start out rough in the forming neural plate (NP) and sharpen over time. However, the mechanisms controlling simultaneous formation of multiple rhombomeres and accuracy in their sizes are unclear. We have developed a stochastic multiscale cell-based model that explicitly incorporates dynamic morphogenetic changes (i.e. convergent-extension of the NP), multiple morphogens, and gene regulatory networks to investigate the formation of rhombomeres and their corresponding boundaries in the zebrafish hindbrain. During pattern initiation, the short-range signal, fibroblast growth factor (FGF), works together with the longer-range morphogen, retinoic acid (RA), to specify all of these boundaries and maintain accurately-sized segments with sharp boundaries. At later stages of patterning, we show a nonlinear change in the shape of rhombomeres with rapid left-right narrowing of the NP followed by slower dynamics. Rapid initial convergence improves boundary sharpness and segment size by regulating cell sorting and cell fate both independently and coordinately. Overall, multiple morphogens and tissue dynamics synergize to regulate the sizes and boundaries of multiple segments during development. Author SummaryIn segmental pattern formation, chemical gradients control gene expression in a concentration-dependent manner to specify distinct gene expression domains. Despite the stochasticity inherent to such biological processes, precise and accurate borders form between segmental gene expression domains. Previous work has revealed synergy between gene regulation and cell sorting in sharpening borders that are initially rough. However, it is still poorly understood how size and boundary sharpness of multiple segments are regulated in a tissue that changes dramatically in its morphology as the embryo develops. Here we develop a stochastic multiscale cell-base model to investigate these questions. Two novel strategies synergize to promote accurate segment formation, a combination of long- and short-range morphogens plus rapid tissue convergence, with one responsible for pattern initiation and the other enabling pattern refinement.

developmental biology↗

bioRxiv2020

Lmo7a Coordinates Neural Crest Migration and Lineage Specification by Regulating Cell Adhesion Dynamics

Cell migration requires dynamic regulation of cell-cell signaling and cell adhesion. Neural crest (NC) cells are highly migratory cells, which undergo an epithelial-mesenchymal transition (EMT) to leave the neural epithelium and migrate throughout the body to give rise to many different derivatives. We have identified a Lim-domain only (Lmo) protein, Lmo7a, expressed in early NC cells that controls both actin cytoskeletal dynamics and Wnt signaling during NC migration. In embryos deficient in Lmo7a, many NC cells fail to migrate away from the dorsal midline, and form aggregates. Unlike the majority of NC cells that appear to migrate normally, cells that aggregate in Lmo7a-deficient embryos mislocalize paxillin (Pxn) and have reduced levels of phosphorylated focal adhesion kinase (pFAK). Lmo7a loss-of-function also disrupts canonical Wnt signaling such that after the onset of NC cell migration, Wnt responses and nuclear {beta}-catenin levels increase in the cells that aggregate. However, this increase in Wnt signaling appears secondary to the defect in migration. Similar to mutants for other Wnt regulators in NC cells, the NC cells in Lmo7a-deficient aggregates exhibit gene expression signatures of pigment cell progenitors, but also express markers of Schwann cell progenitors, suggesting a role for Lmo7a in pigment-glial specification. We propose that Lmo7a modulates cell adhesion to facilitate both robust NC cell migration and a subset of lineage decisions.

developmental biology↗

bioRxiv2017

Cell-type heterogeneity in the zebrafish olfactory placode is generated from progenitors within preplacodal ectoderm

Vertebrate olfactory placodes consists of a variety of neuronal populations, which are thought to have distinct embryonic origins. In the zebrafish, while ciliated sensory neurons arise from preplacodal ectoderm (PPE), previous lineage tracing studies suggest that both Gonadotropin releasing hormone 3 (Gnrh3) and microvillous sensory neurons derive from cranial neural crest (CNC). We find that the expression of Islet1/2 is restricted to Gnrh3 neurons associated with the olfactory placode. Unexpectedly, however, we find no change in Islet1/2+ cell numbers in sox10 mutant embryos, calling into question their CNC origin. Lineage reconstruction based on backtracking in time-lapse confocal datasets, and confirmed by photoconversion experiments, reveals that Gnrh3 neurons derive from the anterior/medial PPE. Similarly, all of the microvillous sensory neurons we have traced arise from preplacodal progenitors. Our results suggest that rather than originating from separate ectodermal populations, cell-type heterogeneity is generated from overlapping pools of progenitors within the preplacodal ectoderm.

developmental biology↗