Biology subjects

McKinney, M. C.

Publications and source records attributed to McKinney, M. C..

13 recordsLinked to original sources

bioRxiv2026

Loss of polr1c disrupts myelination in a zebrafish model of POLR1C-associated disease

Pathogenic variants in POLR1C, which encodes a shared subunit of RNA Polymerases (Pols) I and III, cause Treacher Collins syndrome (TCS) and POLR3-related leukodystrophy. While Pol I and Tp53-dependent mechanisms have been implicated in the pathogenesis of TCS, the basis of hypomyelination in POLR1C-associated POLR3-related leukodystrophy remains incompletely understood. Here, we show that polr1c mutant zebrafish exhibit reduced myelination in addition to previously described craniofacial anomalies. Oligodendrocyte precursor cells exhibit increased activation of the Tp53 pathway; however, these cells do not undergo apoptosis. Consistent with this finding, tp53 inhibition reduces cell death in polr1c mutants but fails to restore myelination, indicating that myelination deficits are not driven by Tp53-dependent progenitor loss in this model. polr1c mutants also exhibit reduced rRNA transcription by Pol I and reduced expression of some Pol III-transcribed tRNAs. Altogether, these data indicate distinct tissue-specific responses to polr1c deficiency and suggest persistent impairment of rRNA transcription contributes to deficient myelin development. These findings expand the developmental consequences of polr1c loss and advance our understanding of the molecular basis of POLR1C-associated diseases.

developmental biology↗

bioRxiv2026

Minicollagen expression dynamics reveal a transcriptional program for cnidogenesis in the sea anemone Nematostella vectensis

Cnidae are explosive harpoon-like organelles localized within stinging cells, or cnidocytes, of the phylum Cnidaria (jellyfish, hydroids, sea anemones, and corals). These unique Golgi-derived vesicular structures define the phylum and are prominent examples of an evolutionary cellular novelty. While recent studies have focused on the developmental specification and regulation of cnidocytes more broadly, less is understood about gene expression patterns, structural variations, and toxin repertoires within distinct cnidae subtypes. Here, we determine the transcriptional profile of two major cnidae subtypes in the sea anemone Nematostella vectensis, nematocytes and spirocytes, using the cnidae-specific structural family of proteins called minicollagens. We first define the in vivo expression patterns for three known and three uncharacterized minicollagen orthologs. We show that four minicollagens are broadly expressed throughout ectodermal cnidocytes in developing larvae and primary polyps while two others are restricted to tentacular cnidocytes. Leveraging whole adult scRNA-seq data and two novel transgenic reporter lines, we then demonstrate that the tentacle-restricted cnidocytes are developing spirocytes that are distinguished by expression of the minicollagen NvNcol5. To deepen our analysis of cnidocyte gene expression, we used a customized RNA-FACS-seq pipeline to determine global transcriptional differences between these two subtypes. This approach identified a suite of differentially expressed genes, illuminating spatial and temporal gene expression dynamics across both developing nematocytes and spirocytes. Altogether, our experiments provide fundamental and novel insights into the specialization of cnidarian stinging cells while establishing a rich set of resources for further investigation.

evolutionary biology↗

bioRxiv2026

Nucleolar Dynamics During Oogenesis

Ribosome biogenesis is a conserved and highly regulated process that starts in the nucleolus, a membrane-less multi-phase organelle. Although the architecture of the nucleolus is known to change due to perturbations, how nucleolar organization is modulated during physiological processes to meet changing translational demands remains unclear. Here, we use zebrafish oogenesis as a developmental context requiring a rapid expansion of translational capacity to investigate the regulation of nucleolar architecture. We show nucleoli undergo coordinated changes in number, size, subnuclear localization, and layering throughout oogenesis. We further demonstrate that nucleoli form around extrachromosomal DNA circles that contain the rDNA locus. Notably, mouse oocytes undergo similar developmental changes in nucleolar layering and phase organization, indicating that remodeling of nucleolar condensates is a conserved feature of oogenesis. These findings reveal previously unexplored regulation of nucleolar architecture as developmental adaptations to changing biosynthetic needs.

developmental biology↗

bioRxiv2026

A versatile cryopreservation method for peri-gastrulation squamate embryos optimised using the veiled chameleon (C. calyptratus)

Stem cell technologies have become a vital component of conservation efforts around the globe. Biobanks and pluripotent stem cell lines help to ensure species and their genetic diversity are preserved. These efforts have however, focussed mostly on mammals and birds, and the cryopreservation protocols for embryos and cells were developed decades ago laying the basis for artificial reproductive techniques for species conservation. With over 20% of non-avian reptile species facing extinction, it is imperative to establish protocols for reptiles to ensure species preservation and also to facilitate the establishment of new reptile model organisms to match the standard of mammals. Here, we have generated a cryopreservation method for preserving early gastrulating veiled chameleon embryos as a representative squamate species. To this end, we first developed a tissue culture method for maintaining cells extracted from peri-gastrulation chameleon embryos and then tested different cryopreservation methods altering the concentration of the penetrating cryoprotectant DMSO and assessing the effect of the addition of non-penetrating cryoprotectants Trehalose and Sucrose. We then optimised a protocol for whole embryo vitrification in 20% DMSO with added Trehalose or Sucrose that can easily be adapted for fieldwork. Taken together, our method not only provides a protocol for conservation efforts but also lays the basis for mechanistic studies of early squamate embryo development by enabling cryopreservation of whole embryos in a fieldwork setting, which facilitates their live transport back to a laboratory for functional experiments or molecular analyses.

developmental biology↗

bioRxiv2024

Comparative Analysis of Neural Crest Development in the Chick and Mouse

A core framework of the gene regulatory network (GRN) governing neural crest (NC) cell development has been generated by integrating separate inputs from diverse model organisms rather than direct comparison. This has limited insights into the diversity of genes in the NC cell GRN and extent of conservation of newly identified transcriptional signatures in cell differentiation and invasion. Here, we address this by leveraging the strengths and accessibility of the avian embryo to precise developmental staging by egg incubation and use an integrated analysis of chick (HH13) and mouse (E9.5) embryo tissue samples collected during NC cell migration into pharyngeal arches 1-2 (PA1 and PA2). We successfully identify a cluster of NC cells containing both mouse and chick cells that share expression of Lmo4, Tfap2B, Sox10, and Twist1, and distinct genes that lack known conserved roles in NC. Importantly, we discovered a cluster of cells exhibiting a conserved transcriptional signature associated with the NC cell migratory wavefront in both mouse and chick, including KAZALD1, BAMBI, DES, and GPC3. We confirm their expression is restricted to leader mouse NCs by multiplexed FISH. Together, these data offer novel insights into the transcriptional programs that underlie NC cell migration and establish the foundation for future comparative functional analyses.

developmental biology↗

bioRxiv2024

Cell extrusion - a novel mechanism driving neural crest cell delamination

Neural crest cells (NCC) comprise a heterogeneous population of cells with variable potency, that contribute to nearly every tissue and organ system throughout the body. Considered unique to vertebrates, NCC are transiently generated within the dorsolateral region of the neural plate or neural tube, during neurulation. Their delamination and migration are crucial events in embryo development as the differentiation of NCC is heavily influenced by their final resting locations. Previous work in avian and aquatic species has shown that NCC delaminate via an epithelial-mesenchymal transition (EMT), which transforms these stem and progenitor cells from static polarized epithelial cells into migratory mesenchymal cells with fluid front and back polarity. However, the cellular and molecular drivers facilitating NCC delamination in mammals are poorly understood. We performed live timelapse imaging of NCC delamination in mouse embryos and discovered a group of cells that exit the neuroepithelium as isolated round cells, which then halt for a short period prior to acquiring the mesenchymal migratory morphology classically associated with most delaminating NCC. High magnification imaging and protein localization analyses of the cytoskeleton, together with measurements of pressure and tension of delaminating NCC and neighboring neuroepithelial cells, revealed these round NCC are extruded from the neuroepithelium prior to completion of EMT. Furthermore, we demonstrate that cranial NCC are extruded through activation of the mechanosensitive ion channel, PIEZO1, a key regulator of the live cell extrusion pathway, revealing a new role for PIEZO1 in neural crest cell development. Our results elucidating the cellular and molecular dynamics orchestrating NCC delamination support a model in which high pressure and tension in the neuroepithelium results in activation of the live cell extrusion pathway and delamination of a subpopulation of NCC in parallel with EMT. This model has broad implications for our understanding of cell delamination in development and disease.

developmental biology↗

bioRxiv2024

Interpretable deep learning reveals the sequence rules of Hippo signaling

The response to signaling pathways is highly context-specific, and identifying the transcription factors and mechanisms that are responsible is very challenging. Using the Hippo pathway in mouse trophoblast stem cells as a model, we show here that this information is encoded in cis-regulatory sequences and can be learned from high-resolution binding data of signaling transcription factors. Using interpretable deep learning, we show that the binding levels of TEAD4 and YAP1 are enhanced in a distance-dependent manner by cell type-specific transcription factors, including TFAP2C. We also discovered that strictly spaced Tead double motifs are widespread highly active canonical response elements that mediate cooperativity by promoting labile TEAD4 protein-protein interactions on DNA. These syntax rules and mechanisms apply genome-wide and allow us to predict how small sequence changes alter the activity of enhancers in vivo. This illustrates the power of interpretable deep learning to decode canonical and cell type-specific sequence rules of signaling pathways. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=183 SRC="FIGDIR/small/580842v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@1ee22e9org.highwire.dtl.DTLVardef@1362c82org.highwire.dtl.DTLVardef@1a42512org.highwire.dtl.DTLVardef@17dd619_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2024

A new genetically tractable non-vertebrate system to study complete camera-type eye regeneration

Camera-type eyes are complex sensory organs susceptible to irreversible damage. Their repair is difficult to study due to the paucity of camera-type eye regeneration models. Identifying a genetically tractable organism with the ability to fully regenerate complete camera-type eyes would help overcome this difficulty. Here, we introduce the apple snail Pomacea canaliculata, capable of full regeneration of camera-type eyes even after complete resection. We defined anatomical components of P. canaliculata eyes and genes expressed during crucial steps of their regeneration. By exploiting the unique features of this organism, we successfully established the first stable mutant lines in apple snails. Our studies revealed that, akin to humans, pax6 is indispensable for eye development in apple snails, establishing this as a research organism to unravel the mechanisms of camera-type eye regeneration. This work expands our understanding of complex sensory organ regeneration and offers new ways to explore this process.

developmental biology↗

bioRxiv2024

Starvation resistant cavefish reveal conserved mechanisms of starvation-induced hepatic lipotoxicity

Starvation causes the accumulation of lipid droplets in the liver, a somewhat counterintuitive phenomenon that is nevertheless conserved from flies to humans. Much like fatty liver resulting from overfeeding, hepatic lipid accumulation (steatosis) during undernourishment can lead to lipotoxicity and atrophy of the liver. Here, we found that while surface populations of Astyanax mexicanus undergo this evolutionarily conserved response to starvation, the starvation-resistant cavefish larvae of the same species do not display an accumulation of lipid droplets upon starvation. Moreover, cavefish are resistant to liver atrophy during starvation, providing a unique system to explore strategies for liver protection. Using comparative transcriptomics between zebrafish, surface fish, and cavefish, we identified the fatty acid transporter slc27a2a/fatp2 to be correlated with the development of fatty liver. Pharmacological inhibition of slc27a2a in zebrafish rescues steatosis and atrophy of the liver upon starvation. Further, down-regulation of FATP2 in drosophila larvae inhibits the development of starvation-induced steatosis, suggesting the evolutionary conserved importance of the gene in regulating fatty liver upon nutrition deprivation. Overall, our study identifies a conserved, druggable target to protect the liver from atrophy during starvation. One-Sentence SummaryCavefish evolved protection from starvation-induced liver damage through reduction of fatty acid uptake regulated by FATP2, a mechanism conserved through 400 million years of animal evolution.

cell biology↗

bioRxiv2023

Graded FGF activity patterns distinct cell types within the apical sensory organ of the sea anemone Nematostella vectensis

Bilaterian animals have evolved complex sensory organs comprised of distinct cell types that function coordinately to sense the environment. Each sensory unit has a defined architecture built from component cell types, including sensory cells, non-sensory support cells, and dedicated sensory neurons. Whether this characteristic cellular composition is present in the sensory organs of non-bilaterian animals is unknown. Here, we interrogate the cell type composition and gene regulatory networks controlling development of the larval apical sensory organ in the sea anemone Nematostella vectensis. Using single cell RNA sequencing and imaging approaches, we reveal two unique cell types in the Nematostella apical sensory organ, GABAergic sensory cells and a putative non-sensory support cell population. Further, we identify the paired-like (PRD) homeodomain gene prd146 as a specific sensory cell marker and show that Prd146+ sensory cells become post-mitotic after gastrulation. Genetic loss of function approaches show that Prd146 is essential for apical sensory organ development. Using a candidate gene knockdown approach, we place prd146 downstream of FGF signaling in the apical sensory organ gene regulatory network. Further, we demonstrate that an aboral FGF activity gradient coordinately regulates the specification of both sensory and support cells. Collectively, these experiments define the genetic basis for apical sensory organ development in a non-bilaterian animal and reveal an unanticipated degree of complexity in a prototypic sensory structure.

developmental biology↗

bioRxiv2022

rRNA transcription is integral to liquid-liquid phase separation and maintenance of nucleolar structure

Beginning with transcription of ribosomal RNA (rRNA) by RNA Polymerase (Pol) I in the nucleolus, ribosome biogenesis is intimately tied to cell growth and proliferation. Perturbation of ribosome biogenesis has been previously shown to affect nucleolar structure, yet the underlying mechanism is unknown. We generated loss-of-function mouse mutants of Pol I subunits, Polr1a, Polr1b, Polr1c and Polr1d, thereby genetically inhibiting rRNA transcription and ribosome biogenesis. Pol I mutant embryos are preimplantation lethal and have fewer nucleoli. Using hiPSCs triple labeled for the three nucleolar compartments, we observe two phenotypes upon Pol I inhibition: a single condensed nucleolus, and fragmented nucleoli. We find that when rRNA transcription is inhibited, the viscosity of the granular compartment of the nucleolus is increased disrupting its liquid-liquid phase separation properties, which results in a condensed nucleolus. Taken together, our data suggests that Pol I function and rRNA transcription are required for maintaining nucleolar structure and integrity.

developmental biology↗

bioRxiv2019

Head Mesoderm Tissue Growth, Dynamics and Neural Crest Cell Migration

Vertebrate head morphogenesis involves orchestrated cell growth and tissue movements of the mesoderm and neural crest to form the distinct craniofacial pattern. To better understand structural birth defects, it is important that we learn how these processes are controlled. Here, we examine this question during chick head morphogenesis using time-lapse imaging, computational modeling, and experiment. We find that head mesodermal cells are inherently dynamic in culture and alter cell behaviors in the presence of either ectoderm or neural crest cells. Mesodermal cells in vivo display large-scale whirling motions that rapidly transition to lateral, directed movements after neural crest cells emerge. Computer model simulations predict distinct changes in neural crest migration as the spatio-temporal growth profile of the mesoderm is varied. BrdU-labeling and photoconversion combined with cell density measurements then reveal non-uniform mesoderm growth in space and time. Chemical inhibition of head mesoderm proliferation or ablation of premigratory neural crest alters mesoderm growth and neural crest migration, implying a dynamic feedback between tissue growth and neural crest cell signaling to confer robustness to the system.\n\nSummary StatementDynamic feedback between tissue growth and neural crest cell migration ensures robust neural crest stream formation and head morphogenesis shown by time-lapse microscopy, mathematical modeling and embryo perturbations.

developmental biology↗

bioRxiv2019

Neural crest cells bulldoze through the microenvironment using Aquaporin-1 to stabilize filopodia

Neural crest migration requires cells to move through an environment filled with dense extracellular matrix and mesoderm to reach targets throughout the vertebrate embryo. Here, we use high-resolution microscopy, computational modeling, and in vitro and in vivo cell invasion assays to investigate the function of Aquaporin-1 (AQP-1) signaling. We find that migrating lead cranial neural crest cells express AQP-1 mRNA and protein, implicating a biological role for water channel protein function during invasion. Differential AQP-1 levels affect neural crest cell speed, direction, and the length and stability of cell filopodia. Further, AQP-1 enhances matrix metalloprotease (MMP) activity and colocalizes with phosphorylated focal adhesion kinases (pFAK). Co-localization of AQP-1 expression with EphB guidance receptors in the same migrating neural crest cells raises novel implications for the concept of guided bulldozing by lead cells during migration.

developmental biology↗