Biology subjects

Zilova, L.

Publications and source records attributed to Zilova, L..

6 recordsLinked to original sources

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↗

bioRxiv2025

Extrinsic polarity cues control lamination versus cluster-based organisation in vertebrate retinal development

1.Photosensitive organs are essential for most animals to perceive and respond to their environment. While the gene regulatory networks establishing retinal identity are deeply conserved across metazoans (reviewed in Gehring, 2012; Vopalensky & Kozmik, 2009; Hahn et al., 2023), the retinal architecture varies widely--from invertebrate compound eyes to vertebrate camera-type eyes (Lamb et al., 2007; Schwab, 2017; Arendt & Wittbrodt, 2001). Despite this morphological diversity, early eye anlagen in both, invertebrates and vertebrates, share an initial pseudo-stratified epithelial organization (Weasner & Kumar, 2022; Randlett et al., 2010; Das et al., 2003; Kitambi & Malicki, 2008), which is maintained and elaborated into multi-layered retinae in vertebrates. In contrast, the invertebrate neuroepithelium is re-organized as ommatidia develop. Laminar organisation of the vertebrate retina appears to be a consequence of initial polarisation of the retinal neuroepithelium. This is, however, challenging to test in the organismal context. To address the plasticity of retinal architecture and the impact of epithelial polarity on the structuring of retinal tissue, we take advantage of retinal organoids derived from medaka (Oryzias latipes) (Zilova et al., 2021) that allow to modulate polarity cues and test their impact on the level of epithelialization and structural organisation of the forming retina. We show that under specific culture conditions, medaka retinal organoids undergo a striking morphological switch depending on the level of apico-basal polarity imposed. When polarity cues are continuously provided, a laminated retinal epithelium is established in the organoid. The absence of polarity cues results in the formation of horizontal cellular clusters containing the retinal cell types, which form the vertical retinal column in the developing embryo. We demonstrate that the emergence of this alternative retinal architecture is associated with a loss of epithelial polarity, notably the absence of extracellular matrix (ECM) components, such as laminin, which efficiently rescues lamination. Our findings indicate that tissue-level polarization and lamination in vertebrate retinae require specific extrinsic cues, and that in their absence, differentiating retinal cell types self-organize into structurally distinct, retinal units. This reveals an unexpected plasticity in vertebrate retinal development and indicates a potential for alternative modes of retinal patterning. O_FIG O_LINKSMALLFIG WIDTH=155 HEIGHT=200 SRC="FIGDIR/small/688026v1_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@100bb1eorg.highwire.dtl.DTLVardef@fceb9borg.highwire.dtl.DTLVardef@d036a7org.highwire.dtl.DTLVardef@1b41483_HPS_FORMAT_FIGEXP M_FIG C_FIG Retinal cells in medaka organoids adopt either a continuous layered epithelium when supported by laminin or a unit-based, ommatidia-like organization when epithelial continuity is lost. This dual outcome suggests that epithelial integrity represents a branching point between vertebrate and invertebrate strategies of retinal patterning, providing an experimental system to replay alternative evolutionary trajectories of eye design.

developmental biology↗

bioRxiv2025

Inverted Assembly of the Lens Within Ocular Organoids Reveals Alternate Paths to Ocular Morphogenesis

The eye is a complex organ composed of two main structures - the retina and the lens. It forms by the invagination of the lens forming head surface ectoderm embedding into the forming optic cup. This "outside-in" mode of morphogenesis ensures that the light focusing lens is positioned centrally inside of the eye in the highly constrained environment of the developing embryo. Advances in stem cell biology in the last decade introduced organoids as model to study organogenesis under normal and diseased conditions. However, even though strikingly similar at some points, it remained elusive to which extend the generation of individual structural features in organoids recapitulates in vivo organogenesis. Here we describe the generation of fish ocular organoids composed of both, lens and retina, using pluripotent embryonic cells from medaka (Oryzias latipes). Formation of the organoid lens followed the key molecular features of the process in vivo, including the establishment of lens progenitor cells and their subsequent differentiation into lens fiber cells. In a process dependent on the coordinated activity of BMP and FGF signaling, lens formation in ocular organoids was marked by the expression of key genes implicated in organismal lens development. Despite adhering to the basic molecular machinery of lens formation in vivo, the morphogenesis into a spherical lens followed an "inside-out" mode. Lens progenitor cells were initially established and differentiated into a spherical lens directly inside of the retina. Subsequent displacement of the lens from the center of the organoid towards its surface ultimately led to the formation of a cup-like shaped retina with a centrally positioned lens. Our study highlights that the self-organization of the organoid can favor routes that were not selected for in the developing embryo. Those routes can lead to an alternative, though highly similar outcome with the respect to achieving specific structural features in an unconstrained, embryo-free environment.

developmental biology↗

bioRxiv2025

Deep learning predicts tissue outcomes in retinal organoids

Retinal organoids have become important models for studying development and disease, yet stochastic heterogeneity in the formation of cell types, tissues, and phenotypes remains a major challenge. This limits our ability to precisely experimentally address the early developmental trajectories towards these outcomes. Here, we utilize deep learning to predict the differentiation path and resulting tissues in retinal organoids well before they become visually discernible. Our approach effectively bypasses the challenge of organoid-related heterogeneity in tissue formation. For this, we acquired a high-resolution time-lapse imaging dataset comprising about 1,000 organoids and over 100,000 images enabling precise temporal tracking of organoid development. By combining expert annotations with advanced image analysis of organoid morphology, we characterized the heterogeneity of the retinal pigmented epithelium (RPE) and lens tissues, as well as global organoid morphologies over time. Using this training set, our deep learning approach accurately predicts the emergence and size of RPE and lens tissue formation on an organoid-by-organoid basis at early developmental stages, refining our understanding of when early lineage decisions are made. This approach advances knowledge of tissue and phenotype decision-making in organoid development and can inform the design of similar predictive platforms for other organoid systems, paving the way for more standardized and reproducible organoid research. Finally, it provides a direct focus on early developmental time points for in-depth molecular analyses, alleviated from confounding effects of heterogeneity.

developmental biology↗

bioRxiv2023

Visualisation of gene expression within the context of tissues: an X-ray computed tomography-based multimodal approach

The development of an organism is orchestrated by the spatial and temporal expression of genes. Accurate visualisation of gene expression patterns in the context of the surrounding tissues offers a glimpse into the mechanisms that drive morphogenesis. We developed correlative light-sheet fluorescence microscopy and X-ray computed tomography approach to map gene expression patterns to the whole organisms 3D anatomy at cellular resolution. We show that this multimodal approach is applicable to gene expression visualised by protein-specific antibodies and fluorescence RNA in situ hybridisation, offering a detailed understanding of individual phenotypic variations in model organisms. Furthermore, the approach provides a unique possibility to identify tissues together with their 3D cellular and molecular composition in anatomically less-defined in vitro models, such as organoids. We anticipate that the visual and quantitative insights into the 3D distribution of gene expression within tissue architecture, by the multimodal approach developed here, will be equally valuable for reference atlases of model organisms development, as well as for comprehensive screens and morphogenesis studies of in vitro models.

developmental biology↗

bioRxiv2021

Fish primary embryonic stem cells self-assemble into retinal tissue mirroring in vivo early eye development

Organoids derived from pluripotent stem cells promise the solution to current challenges in basic and biomedical research. Further progress and widespread applications are however limited by long developmental time, variable success, and lack of direct comparison to an in vivo reference. To overcome those limitations, we derived organoids from rapidly developing teleosts. We demonstrate how primary embryonic stem cells from zebrafish and medaka efficiently self-organize into anterior neural structures, particularly retina. Within four days, blastula-stage cell aggregates reproducibly execute key steps of eye development: retinal specification, morphogenesis and differentiation. The number of aggregated cells as well as genetic factors crucially impacted upon the concomitant morphological changes that were intriguingly reflecting the in vivo situation. High reproducibility and rapid development of fish-derived organoids in combination with advanced genome editing techniques immediately allow addressing aspects of development and disease, and systematically probing the impact of the physical environment on morphogenesis and differentiation.

developmental biology↗