Biology subjects

Farrell, J. A.

Publications and source records attributed to Farrell, J. A..

11 recordsLinked to original sources

bioRxiv2025

Developmental regulation of intestinal best4+ cells

best4+/CFTR-high expressing cells are a recently described intestinal epithelial cell type potentially altered in inflammatory bowel disease and colorectal cancer. However, their developmental origin, developmental regulation, and functions remain undefined. This study identifies their conserved transcriptional program and uses zebrafish to dissect their developmental regulation in vivo. Lineage tracing identified that best4+ cells arise from atoh1b+ secretory progenitors. We identify that Notch signaling, mediated by dll4, specifies best4+ cells at the expense of enterochromaffin cells. Downstream of Notch, meis1b confers best4+ cell identity. best4+ cells then exhibit regionalized gene expression, regulated by pbx3a. Additionally, this study demonstrates a system where best4+ cells can be manipulated, observed, and removed in an organismal context. Live imaging and electron microscopy of best4+ cells identified dynamic cellular projections, suggesting a sensory or communicative function. Removal of best4+ cells in vivo eliminated previously proposed functions: they are not required to restore intestinal pH following acidic challenge and do not absorb nutrients. However, we identify region-specific intracellular pH differences that suggest potential functional heterogeneity. Altogether, this study presents a comprehensive description of best4+ cell development from birth to spatial regulation that will be instrumental to understand how best4+ cells change in disease or might be therapeutically manipulated and presents the tools to dissect their function in vivo.

developmental biology↗

bioRxiv2025

Specialized gas-exchange endothelium of the zebrafish gill

The pulmonary vasculature plays a critical role in gas exchange and in lung pathologies, but it is challenging to observe and experimentally manipulate deep within the lungs of living mammals. Unlike mammalian lungs, externally located zebrafish gills are readily accessible for high-resolution optical imaging and experimental manipulation, suggesting zebrafish might provide an excellent comparative vertebrate model for studying the development and function of gas-exchange organs and the gas-exchange blood vasculature. To characterize their resident cell populations, we performed single-cell RNA sequencing (scRNAseq) on adult zebrafish gills, revealing numerous cell types with transcriptional similarities to those found in mammalian lungs. We uncovered and characterized several different endothelial cell populations, including distinct clusters of arterial endothelial cells and lymphatic endothelial cells. The largest endothelial cell cluster closely resembles Cap2 or "Aerocyte" endothelial cells, a recently discovered unusual mammalian endothelial cell type found exclusively in lung alveoli. Zebrafish aerocytes localize to the analogous gas-exchange structures in fish, the highly vascularized gill lamellae. We use confocal and super-resolution imaging of transgenic and hybridization chain reaction-probed zebrafish, array tomography, and focused ion beam scanning electron microscopy to carry out a detailed and comprehensive characterization of gill aerocytes including 3-D ultrastructural reconstruction of one of these cells, showing that as in mammals these cells are closely associated with gas-exchange epithelia and that they possess unique properties that may help facilitate their gas-exchange function. Together, our findings help establish a new, experimentally accessible comparative vertebrate model for studying the gas-exchange blood vasculature.

developmental biology↗

bioRxiv2025

prdm1a drives a fate switch between hair cells of different mechanosensory organs

Vertebrate mechanosensory hair cells (HCs) in the ear detect sound and gravitational forces. Additionally, fish have homologous lateral line HCs in the skin that detect water vibrations for orientation and predator avoidance. HCs in fish and other non-mammalian vertebrates regenerate to restore function after damage, but mammalian HCs lack this ability, causing deafness and vestibular defects. Experimental attempts at regeneration in mice result in incomplete differentiation of immature HCs. Despite differences in regeneration, the gene regulatory networks (GRNs) driving HC maturation are highly similar across vertebrates. Here, we show that the transcription factor prdm1a plays a key role in the HC fate GRN in the zebrafish lateral line. Mutating prdm1a respecifies lateral line HCs into ear HCs, altering morphology and transcriptome. Understanding how transcription factors control diverse HC fates in zebrafish is crucial for understanding the yet unsolved regeneration of diverse HCs in mammalian ears to restore hearing and balance.

developmental biology↗

bioRxiv2025

Anatomical and Molecular Characterization of the Zebrafish Meninges

The meninges are a set of connective tissue layers that surround the central nervous system, protecting the brain from mechanical shock, supporting its buoyancy, guarding it from infection and injury, and maintaining brain homeostasis. Despite their critical role, the molecular identity, developmental origins, and functional properties of the cell types populating the meninges remain poorly characterized. This is in large part due to lack of cell type specific markers and difficulty in visualizing and studying these structures through the thick mammalian skull. Here, we show that the zebrafish, a genetically and experimentally accessible vertebrate, possesses an easily imaged mammalian-like meninges. Anatomical and cellular characterization of its composition via histology, electron microscopy, and confocal imaging shows that the adult zebrafish possesses complex multilayered meninges with double-layered dura mater and intricate leptomeningeal layers. Using single cell transcriptomics, we define the molecular identities of meningeal cell populations, including a unique ependymin (epd)-expressing cell population that constitutes the major cellular component of the leptomeningeal barrier and is essential for brain development and survival. These findings support the use of zebrafish as a useful comparative model for studying the meninges, provide a foundational description for future zebrafish meningeal research, and identify a new Leptomeningeal Barrier Cell that serves as the primary epithelial cell component of the leptomeninges.

developmental biology↗

bioRxiv2023

Gene module reconstruction elucidates cellular differentiation processes and the regulatory logic of specialized secretion

During differentiation, cells become structurally and functionally specialized, but comprehensive views of the underlying remodeling processes are elusive. Here, we leverage scRNA-seq developmental trajectories to reconstruct differentiation using two secretory tissues as a model system - the zebrafish notochord and hatching gland. First, we present an approach to integrate expression and functional similarities for gene module identification, revealing dozens of gene modules representing known and newly associated differentiation processes and their temporal ordering. Second, we focused on the unfolded protein response (UPR) transducer module to study how general versus cell-type specific secretory functions are regulated. By profiling loss- and gain-of-function embryos, we found that the UPR transcription factors creb3l1, creb3l2, and xbp1 are master regulators of a general secretion program. creb3l1/creb3l2 additionally activate an extracellular matrix secretion program, while xbp1 partners with bhlha15 to activate a gland-specific secretion program. Our study offers a multi-source integrated approach for functional gene module identification and illustrates how transcription factors confer general and specialized cellular functions.

developmental biology↗

bioRxiv2023

Germ cells do not progress through spermatogenesis in the infertile zebrafish testis

Vertebrate spermatogonial stem cells maintain sperm production over the lifetime of an animal but fertility declines with age. While morphological studies have greatly informed our understanding of typical spermatogenesis, the molecular and cellular mechanisms underlying spermatogenesis are not yet understood, particularly with respect to the onset of fertility. We used single-cell RNA sequencing to generate a developmental atlas of the zebrafish testis. Using 5 timepoints across the adult life of a zebrafish, we described cellular profiles in the testis during and after fertility. While all germ cell stages of spermatogenesis are detected in testes from fertile adult zebrafish, testes from older infertile males only contained spermatogonia and a reduced population of spermatocytes. These remaining germ cells are transcriptionally distinct from fertile spermatogonia. Immune cells including macrophages and lymphocytes drastically increase in abundance in infertile testes. Our developmental atlas reveals the cellular changes as the testis ages and defines a molecular roadmap for the regulation of male fertility.

developmental biology↗

bioRxiv2023

Single-cell temporal dynamics reveals the relative contributions of transcription and degradation to cell-type specific gene expression in zebrafish embryos

AbstractDuring embryonic development, pluripotent cells assume specialized identities by adopting particular gene expression profiles. However, systematically dissecting the underlying regulation of mRNA transcription and degradation remains a challenge, especially within whole embryos with diverse cellular identities. Here, we collect temporal cellular transcriptomes of zebrafish embryos, and decompose them into their newly-transcribed (zygotic) and pre-existing (maternal) mRNA components by combining single-cell RNA-Seq and metabolic labeling. We introduce kinetic models capable of quantifying regulatory rates of mRNA transcription and degradation within individual cell types during their specification. These reveal different regulatory rates between thousands of genes, and sometimes between cell types, that shape spatio-temporal expression patterns. Transcription drives most cell-type restricted gene expression. However, selective retention of maternal transcripts helps to define the gene expression profiles of germ cells and enveloping layer cells, two of the earliest specified cell-types. Coordination between transcription and degradation restricts expression of maternal-zygotic genes to specific cell types or times, and allows the emergence of spatio-temporal patterns when overall mRNA levels are held relatively constant. Sequence-based analysis links differences in degradation to specific sequence motifs. Our study reveals mRNA transcription and degradation events that control embryonic gene expression, and provides a quantitative approach to study mRNA regulation during a dynamic spatio-temporal response.

genomics↗

bioRxiv2023

Single-cell analysis of shared signatures and transcriptional diversity during zebrafish development

During development, animals generate distinct cell populations with specific identities, functions, and morphologies. We mapped transcriptionally distinct populations across 489,686 cells from 62 stages during wild-type zebrafish embryogenesis and early larval development (3-120 hours post-fertilization). Using these data, we identified the limited catalog of gene expression programs reused across multiple tissues and their cell-type-specific adaptations. We also determined the duration each transcriptional state is present during development and suggest new long-term cycling populations. Focused analyses of non-skeletal muscle and the endoderm identified transcriptional profiles of understudied cell types and subpopulations, including the pneumatic duct, individual intestinal smooth muscle layers, spatially distinct pericyte subpopulations, and homologs of recently discovered human best4+ enterocytes. The transcriptional regulators of these populations remain unknown, so we reconstructed gene expression trajectories to suggest candidates. To enable additional discoveries, we make this comprehensive transcriptional atlas of early zebrafish development available through our website, Daniocell.

developmental biology↗

bioRxiv2023

Differentiation trajectories of the Hydra nervous system reveal transcriptional regulators of neuronal fate

Hydra vulgaris, a cnidarian with a simple nerve net, is an emerging model for developmental, regenerative, and functional neuroscience. Its genetic tractability and capacity for whole-system imaging make it well suited for studying neuron replacement, regeneration, and neural circuit function. Here, we present the most comprehensive molecular and spatial characterization of the H. vulgaris nervous system to date. Using single-cell RNA sequencing, we identified eight neuron types, each defined by distinct neuropeptide expression, and further resolved these into fifteen transcriptionally distinct subtypes with unique spatial distributions and morphologies. To investigate the gene regulatory networks underlying neuronal differentiation, we applied trajectory inference, identified key transcription factors, and performed ATAC-seq on sorted neurons to map chromatin accessibility. All datasets are available through an interactive, user-friendly web portal to support broad use by the research community. Together, these resources provide a foundation for uncovering molecular mechanisms that govern nervous system development, homeostasis, and regeneration in H. vulgaris. Summary StatementHydra vulgaris is a model for regenerative and functional neuroscience. This study identifies fifteen neuron subtypes using scRNA-seq, maps spatial distributions, explores regulatory mechanisms, and provides an accessible web portal.

developmental biology↗

bioRxiv2019

Emergence of neuronal diversity during vertebrate brain development

Neurogenesis in the vertebrate brain comprises many steps ranging from the proliferation of progenitors to the differentiation and maturation of neurons. Although these processes are highly regulated, the landscape of transcriptional changes and progenitor identities underlying brain development are poorly characterized. Here, we describe the first developmental single-cell RNA-seq catalog of more than 200,000 zebrafish brain cells encompassing 12 stages from 12 hours post-fertilization to 15 days post-fertilization. We characterize known and novel gene markers for more than 800 clusters across these timepoints. Our results capture the temporal dynamics of multiple neurogenic waves from embryo to larva that expand neuronal diversity from [~]20 cell types at 12 hpf to [~]100 cell types at 15 dpf. We find that most embryonic neural progenitor states are transient and transcriptionally distinct from long-lasting neural progenitors of post-embryonic stages. Furthermore, we reconstruct cell specification trajectories for the retina and hypothalamus, and identify gene expression cascades and novel markers. Our analysis reveal that late-stage retinal neural progenitors transcriptionally overlap cell states observed in the embryo, while hypothalamic neural progenitors become progressively distinct with developmental time. These data provide the first comprehensive single-cell transcriptomic time course for vertebrate brain development and suggest distinct neurogenic regulatory paradigms between different stages and tissues.

developmental biology↗

bioRxiv2018

Stem cell differentiation trajectories in Hydra resolved at single-cell resolution

The adult Hydra polyp continuously renews all of its cells using three separate stem cell populations, but the genetic pathways enabling homeostatic tissue maintenance are not well understood. We used Drop-seq to sequence transcriptomes of 24,985 single Hydra cells and identified the molecular signatures of a broad spectrum of cell states, from stem cells to terminally differentiated cells. We constructed differentiation trajectories for each cell lineage and identified the transcription factors expressed along these trajectories, thus creating a comprehensive molecular map of all developmental lineages in the adult animal. We unexpectedly found that neuron and gland cell differentiation transits through a common progenitor state, suggesting a shared evolutionary history for these secretory cell types. Finally, we have built the first gene expression map of the Hydra nervous system. By producing a comprehensive molecular description of the adult Hydra polyp, we have generated a resource for addressing fundamental questions regarding the evolution of developmental processes and nervous system function.

developmental biology↗