Biology subjects

Aulehla, A.

Publications and source records attributed to Aulehla, A..

12 recordsLinked to original sources

bioRxiv2026

Spatio-temporal visualisation and manipulation of endogenous Yap1 in medaka

A key requirement to address fundamental questions in developmental biology is the ability to visualise and manipulate endogenous signalling proteins in vivo. Here we generate such an enabling experimental tool for the highly conserved Hippo pathway and its transcriptional effector Yap1, which converts mechanical and chemical cues into transcriptional output during embryogenesis and regeneration. We generate and characterise a Yap1- mGreenLantern (Yap1-mGL) endogenous CRISPR/Cas9-mediated knock-in line that enables 4D analysis of Yap1 dynamics in medaka. We resolve Yap1- mGL expression and nuclear/cytoplasmic localisation in real time, revealing the onset of reporter fluorescence before gastrulation, tissue and cell-type specific Yap1-mGL localisation during somitogenesis and injury-induced Yap1 upregulation during larval spinal cord regeneration. Using the nanobody-based degron system, we degrade the Yap1 protein in vivo, recapitulating the genetic loss-of-function mutant phenotype. Combining endogenous tags with nanobody-based degrons offers a powerful approach to visualise and acutely perturb endogenous protein function in vivo.

developmental biology↗

bioRxiv2026

Quantitative mapping of heterochrony to species-specific phenotypes

The genetic program of animal development is conserved, but its rate of execution varies across species. Heterochrony, shifts in the relative timing of developmental events, generates phenotypic variation, but its prevalence and origin are unclear. Here, we compare two vertebrates with a 3-fold difference in developmental rate, zebrafish (Danio rerio) and medaka (Oryzias latipes) and characterize heterochrony with embryo-scale single-cell genomics. We generated an atlas of >1.2M single-cell transcriptomes of medaka from blastula to hatch and developed a new approach to represent medaka development in "zebrafish time", uncovering many cryptic, cell-level timing shifts not predicted by medakas slower development. We confirm a divergent heterochrony in the medaka notochord using in vivo imaging, revealing that coupled acceleration and delay of sister cell types shapes the species-specific morphology of this tissue. Our results point to cell type-specific timing deviations as a reservoir of phenotypic variation and we propose that species-specific developmental rate can emerge from these cell-level differences.

Developmental Biology↗

bioRxiv2026

Cycles upon cycles - Temperature Scaling of Medaka Development

How organisms develop in dynamic environmental conditions is a fundamental question. We asked how day-night temperature cycles impact embryonic axis elongation and segmentation, itself a cyclic process linked to the segmentation clock, using the Japanese rice fish medaka. We developed an unbiased dimensional reduction approach, based on Singular Value Decomposition (SVD), to reliably identify the dynamic modes of segmentation clock oscillations across all temperature conditions. We reveal that the two major dynamic modes show opposite temperature sensitivities: while the temporal oscillation (mode 1) varies strongly with temperature, the spatial phase gradient (mode 2) appears largely temperature invariant. In addition, we found developmental parameters with intermediate, sub-scaled temperature responses, such as axis elongation. We used theoretical modeling to understand how dynamic modes emerge from the underlying local oscillation dynamics and axis elongation. We then exposed embryos to circadian and ultradian temperature cycles to reveal dynamic response patterns of oscillations and axis elongation, and found how these responses are integrated into morphological features. Combined, our theoretical-experimental results support a model in which the dynamic integration of temporal (i.e. segmentation clock related) and spatial (i.e. axis elongation) processes, in particular their sub-scaled temperature response patterns, quantitatively compensate each other to yield a robust, temperature-invariant axis patterning outcome.

developmental biology↗

bioRxiv2026

Heterochrony of axis segmentation underlies extreme morphogenesis in the Japanese eel

Heterochrony is a major mode of vertebrate body plan evolution, yet its molecular and cellular basis remains poorly characterized. Here, we show that axis segmentation in Anguilla japonica--a species that forms 120 vertebrae--is driven by the temporal extension of somitogenesis. This is achieved through the prolonged maintenance of axial progenitors in the tail beyond the hatching stage, coupled to an extreme segment scaling regime that operates under the constraint of minimal axis growth. We identify delayed Hox13 activation and sustained Oct4 expression as molecular signatures of the prolonged segmentation program. Furthermore, we describe two spatially distinct axial progenitor pools that expand stemness in the tail. These findings reveal how the modulation of stemness in time and space drives extreme morphological evolution in vertebrates. One-Sentence SummarySpatiotemporal modulation of stemness drives extreme axis segmentation in the Japanese eel.

developmental biology↗

bioRxiv2026

The limits of scaling in aggregation-driven patterning of cell collectives

Robust development requires maintaining correct proportions as overall size varies. What controls and limits this scaling remains poorly understood due to the complex interplay of mechanical and biochemical embryonic factors. Combining confinement of dissociated embryonic presomitic mesoderm cells with imaging and chemical perturbations, we identified cell aggregation as the initial event in de novo posterior pole formation. Using a continuum model based solely on cell-cell attraction, we quantitatively map how the time available for aggregation-driven patterning limits the maximum scalable system size. Small systems allow rapid, robust pattern scaling, whereas coarsening dynamics substantially delay scaled patterns in large systems. Our experiments quantitatively confirm these predicted scaling regimes. Together, our results reveal a developmental time-size tradeoff governing the scaling of aggregation-driven patterns.

biophysics↗

bioRxiv2024

Multi-species community platform for comparative neuroscience in teleost fish

Studying neural mechanisms in complementary model organisms from different ecological niches in the same animal class can leverage the comparative brain analysis at the cellular level. To advance such a direction, we developed a unified brain atlas platform and specialized tools that allowed us to quantitatively compare neural structures in two teleost larvae, medaka (Oryzias latipes) and zebrafish (Danio rerio). Leveraging this quantitative approach we found that most brain regions are similar but some subpopulations are unique in each species. Specifically, we confirmed the existence of a clear dorsal pallial region in the telencephalon in medaka lacking in zebrafish. Further, our approach allows for extraction of differentially expressed genes in both species, and for quantitative comparison of neural activity at cellular resolution. The web-based and interactive nature of this atlas platform will facilitate the teleost communitys research and its easy extensibility will encourage contributions to its continuous expansion.

neuroscience↗

bioRxiv2024

Nonreciprocal synchronization in embryonic oscillator ensembles

Synchronization of coupled oscillators is a universal phenomenon encountered across different scales and contexts e.g., chemical wave patterns, superconductors and the unison applause we witness in concert halls. The existence of common underlying coupling rules define universality classes, revealing a fundamental sameness between seemingly distinct systems. Identifying rules of synchronization in any particular setting is hence of paramount relevance. Here, we address the coupling rules within an embryonic oscillator ensemble linked to vertebrate embryo body axis segmentation. In vertebrates, the periodic segmentation of the body axis involves synchronized signaling oscillations in cells within the presomitic mesoderm (PSM), from which somites, the pre-vertebrae, form. At the molecular level, it is known that intact Notch-signaling and cell-to-cell contact is required for synchronization between PSM cells. However, an understanding of the coupling rules is still lacking. To identify these, we develop a novel experimental assay that enables direct quantification of synchronization dynamics within mixtures of oscillating cell ensembles, for which the initial input frequency and phase distribution are known. Our results reveal a "winner-takes-it-all" synchronization outcome i.e., the emerging collective rhythm matches one of the input rhythms. Using a combination of theory and experimental validation, we develop a new coupling model, the "Rectified Kuramoto" (ReKu) model, characterized by a phase-dependent, non-reciprocal interaction in the coupling of oscillatory cells. Such non-reciprocal synchronization rules reveal fundamental similarities between embryonic oscillators and a class of collective behaviours seen in neurons and fireflies, where higher level computations are performed and linked to non-reciprocal synchronization.

systems biology↗

bioRxiv2024

Glycolysis-Wnt signaling axis tunes developmental timing of embryo segmentation

The question of how metabolism impacts development is seeing a renaissance [1, 2]. How metabolism exerts instructive signaling functions is one of the central issues that need to be resolved. We tackled this question in the context of mouse embryonic axis segmentation. Previous studies have shown that changes in central carbon metabolism impact Wnt signaling [3-6] and the period of the segmentation clock [7], which controls the timing of axis segmentation. Here, we reveal that glycolysis tunes the segmentation clock period in an anti-correlated manner: higher glycolytic flux slows down the clock, and vice versa. Transcriptome and gene regulatory network analyses identified Wnt signaling and specifically the transcription factor Tcf7l2, previously associated with increased risk for diabetes [8, 9], as potential mechanisms underlying flux-dependent control of the clock period. Critically, we show that deletion of the Wnt antagonist Dkk1 rescued the slow segmentation clock phenotype caused by increased glycolysis, demonstrating that glycolysis instructs Wnt signaling to control the clock period. In addition, we demonstrate metabolic entrainment of the segmentation clock: periodic changes in the levels of glucose or glycolytic sentinel metabolite fructose 1,6-bisphosphate (FBP) synchronize signaling oscillations. Notably, periodic FBP pulses first entrained Wnt signaling oscillations and subsequently Notch signaling oscillations. We hence conclude that metabolic entrainment has an immediate, specific effect on Wnt signaling. Combined, our work identifies a glycolysis-FBP-Wnt signaling axis that tunes developmental timing, highlighting the instructive signaling role of metabolism in embryonic development.

developmental biology↗

bioRxiv2023

Modular control of time and space during vertebrate axis segmentation

How temporal and spatial control of developmental processes are linked remains a fundamental question. Do underlying mechanisms form a single functional unit or are these dissociable modules? We address this question by studying the periodic process of embryonic axis segmentation, using genetic crosses of inbred medaka fish strains representing two species, Oryzias sakaizumii and latipes. Our analysis revealed correlated interspecies differences with regard to the timing of segmentation, the size of segments and of the presomitic mesoderm (PSM), from which segments are periodically formed. We then did interspecies crosses and real-time imaging quantifications, which revealed extensive phenotypic variation in [~]600 F2 embryos. Importantly, while the F2 analysis showed correlated changes of PSM and segment size, these spatial measures were not correlated to the timing of segmentation. This shows that the control of time and space of axis segmentation can, in principle, be decoupled. In line with this finding, we identified, using developmental quantitative trait loci (devQTL) mapping, distinct chromosomal regions linked to either the control of segmentation timing or PSM size. We were able to validate the devQTL findings using a CRISPR/Cas9 loss-of-function approach on several candidate genes in vivo. Combined, this study reveals that a developmental constraint mechanism underlies spatial scaling of axis segmentation, while its spatial and temporal control are dissociable modules. Our findings emphasise the need to reveal the selective constraints linking these modules in the natural environment.

developmental biology↗

bioRxiv2021

Glycolytic flux-signaling controls mouse embryo mesoderm development

How cellular metabolic state impacts cellular programs is a fundamental, unresolved question. Here we investigated how glycolytic flux impacts embryonic development, using presomitic mesoderm (PSM) patterning as the experimental model. First, we identified fructose 1,6-bisphosphate (FBP) as an in vivo sentinel metabolite that mirrors glycolytic flux within PSM cells of post-implantation mouse embryos. We found that medium-supplementation with FBP, but not with other glycolytic metabolites, such as fructose 6-phosphate and 3-phosphoglycerate, impaired mesoderm segmentation. To genetically manipulate glycolytic flux and FBP levels, we generated a mouse model enabling the conditional overexpression of dominant active, cytoplasmic Pfkfb3 (cytoPfkfb3). Overexpression of cytoPfkfb3 indeed led to increased glycolytic flux/FBP levels and caused an impairment of mesoderm segmentation, paralleled by the downregulation of Wnt-signaling, reminiscent of the effects seen upon FBP-supplementation. To probe for mechanisms underlying glycolytic flux-signaling, we performed subcellular proteome analysis and revealed that cytoPfkfb3 overexpression altered subcellular localization of certain proteins, including glycolytic enzymes, in PSM cells. Specifically, we revealed that FBP supplementation caused depletion of Pfkl and Aldoa from the nuclear-soluble fraction. Combined, we propose that FBP functions as a flux-signaling metabolite connecting glycolysis and PSM patterning, potentially through modulating subcellular protein localization.

developmental biology↗

bioRxiv2021

Arnold tongue entrainment reveals dynamicalprinciples of the embryonic segmentation clock

Living systems exhibit an unmatched complexity, due to countless, entangled interactions across scales. Here we aim to understand a complex system, i.e. segmentation timing in mouse embryos, without a reference to these detailed interactions. To this end, we develop a coarse-grained approach, in which theory guides the experimental identification of the segmentation clock entrainment responses. We demonstrate period- and phase-locking of the segmentation clock across a wide range of entrainment parameters, including higher-order coupling. These quantifications allow to derive the phase response curve (PRC) and Arnold tongues of the segmentation clock, revealing its essential dynamical properties. Our results indicate that the somite segmentation clock has characteristics reminiscent of a highly non-linear oscillator close to an infinite period bifurcation and suggests the presence of long-term feedbacks. Combined, this coarse-grained theoretical-experimental approach reveals how we can derive simple, essential features of a highly complex dynamical system, providing precise experimental control over the pace and rhythm of the somite segmentation clock.

developmental biology↗

bioRxiv2021

Endogenous protein tagging in medaka using a simplified CRISPR/Cas9 knock-in approach

The CRISPR/Cas9 system has been used to generate fluorescently labelled fusion proteins by homology directed repair in a variety of species. Despite its revolutionary success, there remains an urgent need for increased simplicity and efficiency of genome editing in research organisms. Here, we establish a simplified, highly efficient and precise strategy for CRISPR/Cas9 mediated endogenous protein tagging in medaka (Oryzias latipes). We use a cloning-free approach that relies on PCR amplified donor fragments containing the fluorescent reporter sequences flanked by short homology arms (30-40bp), a synthetic sgRNA and streptavidin tagged Cas9. We generate six novel knock-in lines with high efficiency of F0 targeting and germline transmission. Whole Genome Sequencing (WGS) results reveal single-copy integration events only at the targeted loci. We provide an initial characterization of these fusion-protein lines, significantly expanding the repertoire of genetic tools available in medaka. In particular, we show that the mScarlet-pcna knock-in line has the potential to serve as an organismal-wide label for proliferative zones and an endogenous cell cycle reporter.

developmental biology↗