Biology subjects

Lewis, V. M.

Publications and source records attributed to Lewis, V. M..

6 recordsLinked to original sources

bioRxiv2024

Early exercise disrupts a pro-repair extracellular matrix program during zebrafish fin regeneration

Mechanical stimulation effects on cell behaviors that restore organ form and function during tissue repair are unresolved. We applied swim flume-mediated exercise during zebrafish caudal fin regeneration to explore mechanical loading impacts on a robust model of organ regeneration. Exercise initiated during but not after blastema establishment compromised fin regeneration, including outgrowth and skeletal pattern. Long-term tracking of fluorescently labeled fibroblasts showed exercise loading disrupted blastemal mesenchyme formation. Transcriptomic profiling and section staining indicated loading reduced an extracellular matrix (ECM) gene expression program, including for hyaluronic acid (HA) synthesis. As with exercise loading, HA synthesis inhibition or blastemal HA depletion impaired blastema formation. We considered if injury-upregulated HA establishes a pro-regenerative environment facilitating mechanotransduction. HA density across the blastema correlated with nuclear localization of the mechanotransducer Yes-associated protein (Yap). Exercise loading or HA depletion decreased nuclear Yap, and culturing primary fin fibroblasts on HA-coated surfaces induced Yap nuclear localization. We conclude early exercise during fin regeneration disrupts expression of an HA-rich ECM supporting Yap-promoted blastema expansion. These findings reveal that fin regeneration is acutely sensitive to the timing and intensity of mechanical loading, underscoring how biomechanical forces integrate with regenerative programs to guide robust tissue repair.

developmental biology↗

bioRxiv2024

Voltage-gated calcium channels generate blastema Ca2+ fluxes restraining zebrafish fin regenerative outgrowth

Adult zebrafish fins regenerate to their original size regardless of damage extent, providing a tractable model of organ size and scale control. Gain-of-function of voltage-gated K+ channels expressed in fibroblast-lineage blastema cells promotes excessive fin outgrowth, leading to a long-finned phenotype. Similarly, inhibition of the Ca2+-dependent phosphatase calcineurin during regeneration causes dramatic fin overgrowth. However, Ca2+ fluxes and their potential origins from dynamic membrane voltages have not been explored or linked to fin size restoration. We used fibroblast-lineage GCaMP imaging of regenerating adult fins to identify widespread, heterogeneous Ca2+ transients in distal blastema cells. Membrane depolarization of isolated regenerating fin fibroblasts triggered Ca2+ spikes dependent on voltage-gated Ca2+ channel activity. Single cell transcriptomics identified the voltage-gated Ca2+ channels cacna1c (L-type channel), cacna1ba (N-type), and cacna1g (T-type) as candidate mediators of fibroblast-lineage Ca2+ signaling. Small molecule inhibition revealed L- and/or N-type voltage-gated Ca2+ channels act during regenerative outgrowth to restore fins to their original scale. Strikingly, cacna1g homozygous mutant zebrafish regenerated extraordinarily long fins due to prolonged outgrowth. The regenerated fins far exceeded their original length but with otherwise normal ray skeletons. Therefore, cacna1g mutants uniquely provide a genetic loss-of-function long-finned model that decouples developmental and regenerative fin outgrowth. Live GCaMP imaging of regenerating fins showed T-type Cacna1g channels enable Ca2+ dynamics in distal fibroblast-lineage blastemal mesenchyme during the outgrowth phase. We conclude "bioelectricity" for fin size control likely entirely reflects voltage-modulated Ca2+ dynamics in fibroblast-lineage blastemal cells that specifically and steadily decelerates outgrowth at a rate tuned to restore the original fin size.

developmental biology↗

bioRxiv2023

The Fraser complex interconnects tissue layers to support basal epidermis and osteoblast integrated morphogenesis underlying fin skeletal patterning

Fraser Syndrome is a rare, multisystemic autosomal recessive disorder characterized by disrupted epithelial-mesenchymal associations upon loss of Fraser Complex genes. Disease manifestation and affected organs are highly variable. Digit malformations such as syndactyly are common but of unclear developmental origins. We explored if zebrafish fraser extracellular matrix complex subunit 1 (fras1) mutants model Fraser Syndrome-associated appendicular skeleton patterning defects. Approximately 10% of fras1 mutants survive to adulthood, displaying striking and varied fin abnormalities, including endochondral bone fusions, ectopic cartilage, and disrupted caudal fin symmetry. The fins of surviving fras1 mutants frequently have fewer and unbranched bony rays. fras1 mutant fins regenerate to their original size but with exacerbated ray branching and fin symmetry defects. Single cell RNA-Seq analysis, in situ hybridizations, and antibody staining show specific Fraser complex expression in the basal epidermis during regenerative outgrowth. Fras1 and Fraser Complex component Frem2 accumulate along the basal side of distal-most basal epidermal cells. Greatly reduced and mislocalized Frem2 accompanies loss of Fras1 in fras1 mutants. The Sonic hedgehog signaling between distal basal epidermis and adjacent mesenchymal pre-osteoblasts that promotes ray branching persists upon Fraser Complex loss. However, fras1 mutant regenerating fins exhibit extensive sub-epidermal blistering associated with a disorganized basal epidermis and adjacent pre-osteoblasts. We propose Fraser Complex-supported tissue layer adhesion enables robust integrated tissue morphogenesis involving the basal epidermis and osteoblasts. Further, we establish zebrafish fin development and regeneration as an accessible model to explore mechanisms of Fraser Syndrome-associated digit defects and Fraser Complex function at epithelial-mesenchymal interfaces.

developmental biology↗

bioRxiv2022

Insulin-like growth factor receptor / mTOR signaling elevates global translation to accelerate zebrafish fin regenerative outgrowth

Zebrafish robustly regenerate fins, including their characteristic bony ray skeleton. Amputation activates intra-ray fibroblasts and dedifferentiates osteoblasts that migrate under a wound epidermis to establish an organized blastema. Coordinated proliferation and re-differentiation across lineages then sustains progressive outgrowth. We generate a single cell transcriptome dataset to characterize regenerative outgrowth and explore coordinated cell behaviors. We computationally identify sub-classes of fibroblast-lineage cells and describe novel markers of intra- and inter-ray fibroblasts and growth-promoting distal blastema cells. A pseudotemporal trajectory and in vivo photoconvertible lineage tracing indicate distal blastemal mesenchyme generates both re-differentiated intra- and inter-ray fibroblasts. Gene expression profiles across this trajectory suggest elevated protein translation in the blastemal mesenchyme state. O-propargyl-puromycin incorporation and small molecule inhibition identify insulin growth factor receptor (IGFR) / mechanistic target of rapamycin kinase (mTOR)-dependent elevated bulk translation in blastemal mesenchyme and differentiating osteoblasts. We test candidate cooperating differentiation factors identified from the osteoblast trajectory, finding IGFR/mTOR signaling expedites glucocorticoid-promoted osteoblast differentiation in vitro. Concordantly, mTOR inhibition slows but does not prevent fin regenerative outgrowth in vivo. IGFR/mTOR may elevate translation in both fibroblast- and osteoblast-lineage cells during the outgrowth phase as a tempo-coordinating rheostat.

developmental biology↗

bioRxiv2021

Development and genetics of red coloration in the zebrafish relative Danio albolineatus

Animal pigment patterns play important roles in behavior and, in many species, red coloration serves as an honest signal of individual quality in mate choice. Among Danio fishes, some species develop erythrophores, pigment cells that contain red ketocarotenoids, whereas other species, like zebrafish (D. rerio) only have yellow xanthophores. Here, we use pearl danio (D. albolineatus) to assess the developmental origin of erythrophores and their mechanisms of differentiation. We show that erythrophores in the fin of D. albolineatus share a common progenitor with xanthophores and maintain plasticity in cell fate even after differentiation. We further identify the predominant ketocarotenoids that confer red coloration to erythrophores and use reverse genetics to pinpoint genes required for the differentiation and maintenance of these cells. Our analyses are a first step towards defining the mechanisms underlying the development of erythrophore-mediated red coloration in Danio and reveal striking parallels with the mechanism of red coloration in birds.

developmental biology↗

bioRxiv2021

Pigment pattern morphospace of Danio fishes: evolutionary diversification and mutational effects

Molecular and cellular mechanisms underlying differences in adult form remain largely unknown. Adult pigment patterns of fishes in the genus Danio, which includes zebrafish, D. rerio, include horizontal stripes, vertical bars, spots and uniform patterns, and provide an outstanding opportunity to identify causes of species level variation in a neural crest derived trait. Yet understanding such variation requires quantitative approaches to assess phenotypes, and such methods have been mostly lacking for pigment patterns. We introduce metrics derived from information theory that describe patterns and pattern variation in Danio fishes. We find that such metrics used singly and in multivariate combinations are suitable for distinguishing general pattern types, and can reveal even subtle phenotypic differences attributable to mutations. Our study provides new tools for analyzing pigment pattern in Danio and potentially other groups, and sets the stage for future analyses of pattern morphospace and its mechanistic underpinnings. Summary statementWe provide quantitative metrics for studying pigment patterns of zebrafish and other species. These metrics are applicable to changes between species as well as impacts of laboratory induced mutations

developmental biology↗