Modeling the extracellular matrix in cell migration and morphogenesis: a guide for the curious biologist.

The extracellular matrix (ECM) is a highly complex structure through which biochemical and mechanical signals are transmitted. In processes of cell migration, the ECM also acts as a scaffold, providing structural support to cells as well as points of potential attachment. Although the ECM is a well-studied structure, its role in many biological processes remains difficult to investigate comprehensively due to its complexity and structural variation within an organism. In tandem with experiments, mathematical models are helpful in refining and testing hypotheses, generating predictions, and exploring conditions outside the scope of experiments. Such models can be combined and calibrated with in vivo and in vitro data to identify critical cell-ECM interactions that drive developmental and homeostatic processes, or the progression of diseases. In this review, we focus on mathematical and computational models of the ECM in processes such as cell migration including cancer metastasis, and in tissue structure and morphogenesis. By highlighting the predictive power of these models, we aim to help bridge the gap between experimental and computational approaches to studying the ECM and to provide guidance on selecting an appropriate model framework to complement corresponding experimental studies.

Hybrid cellular Potts and bead-spring modeling of cells in fibrous extracellular matrix.

The mechanical interaction between cells and the extracellular matrix (ECM) is fundamental to coordinate collective cell behavior in tissues. Relating individual cell-level mechanics to tissue-scale collective behavior is a challenge that cell-based models such as the cellular Potts model (CPM) are well-positioned to address. These models generally represent the ECM with mean-field approaches, which assume substrate homogeneity. This assumption breaks down with fibrous ECM, which has nontrivial structure and mechanics. Here, we extend the CPM with a bead-spring model of ECM fiber networks modeled using molecular dynamics. We model a contractile cell pulling with discrete focal adhesion-like sites on the fiber network and demonstrate agreement with experimental spatiotemporal fiber densification and displacement. We show that at high network cross-linking, contractile cell forces propagate over at least eight cell diameters, decaying with distance with power law exponent n= 0.35 - 0.65 typical of viscoelastic ECMs. Further, we use in silico atomic force microscopy to measure local cell-induced network stiffening consistent with experiments. Our model lays the foundation for investigating how local and long-ranged cell-ECM mechanobiology contributes to multicellular morphogenesis.

The Medaka Inbred Kiyosu-Karlsruhe (MIKK) panel.

BACKGROUND: Unraveling the relationship between genetic variation and phenotypic traits remains a fundamental challenge in biology. Mapping variants underlying complex traits while controlling for confounding environmental factors is often problematic. To address this, we establish a vertebrate genetic resource specifically to allow for robust genotype-to-phenotype investigations. The teleost medaka (Oryzias latipes) is an established genetic model system with a long history of genetic research and a high tolerance to inbreeding from the wild. RESULTS: Here we present the Medaka Inbred Kiyosu-Karlsruhe (MIKK) panel: the first near-isogenic panel of 80 inbred lines in a vertebrate model derived from a wild founder population. Inbred lines provide fixed genomes that are a prerequisite for the replication of studies, studies which vary both the genetics and environment in a controlled manner, and functional testing. The MIKK panel will therefore enable phenotype-to-genotype association studies of complex genetic traits while allowing for careful control of interacting factors, with numerous applications in genetic research, human health, drug development, and fundamental biology. CONCLUSIONS: Here we present a detailed characterization of the genetic variation across the MIKK panel, which provides a rich and unique genetic resource to the community by enabling large-scale experiments for mapping complex traits.

Genomic variations and epigenomic landscape of the Medaka Inbred Kiyosu-Karlsruhe (MIKK) panel.

BACKGROUND: The teleost medaka (Oryzias latipes) is a well-established vertebrate model system, with a long history of genetic research, and multiple high-quality reference genomes available for several inbred strains. Medaka has a high tolerance to inbreeding from the wild, thus allowing one to establish inbred lines from wild founder individuals. RESULTS: We exploit this feature to create an inbred panel resource: the Medaka Inbred Kiyosu-Karlsruhe (MIKK) panel. This panel of 80 near-isogenic inbred lines contains a large amount of genetic variation inherited from the original wild population. We use Oxford Nanopore Technologies (ONT) long read data to further investigate the genomic and epigenomic landscapes of a subset of the MIKK panel. Nanopore sequencing allows us to identify a large variety of high-quality structural variants, and we present results and methods using a pan-genome graph representation of 12 individual medaka lines. This graph-based reference MIKK panel genome reveals novel differences between the MIKK panel lines and standard linear reference genomes. We find additional MIKK panel-specific genomic content that would be missing from linear reference alignment approaches. We are also able to identify and quantify the presence of repeat elements in each of the lines. Finally, we investigate line-specific CpG methylation and performed differential DNA methylation analysis across these 12 lines. CONCLUSIONS: We present a detailed analysis of the MIKK panel genomes using long and short read sequence technologies, creating a MIKK panel-specific pan genome reference dataset allowing for investigation of novel variation types that would be elusive using standard approaches.

Twisting of the zebrafish heart tube during cardiac looping is a tbx5-dependent and tissue-intrinsic process.

Organ laterality refers to the left-right asymmetry in disposition and conformation of internal organs and is established during embryogenesis. The heart is the first organ to display visible left-right asymmetries through its left-sided positioning and rightward looping. Here, we present a new zebrafish loss-of-function allele for tbx5a, which displays defective rightward cardiac looping morphogenesis. By mapping individual cardiomyocyte behavior during cardiac looping, we establish that ventricular and atrial cardiomyocytes rearrange in distinct directions. As a consequence, the cardiac chambers twist around the atrioventricular canal resulting in torsion of the heart tube, which is compromised in tbx5a mutants. Pharmacological treatment and ex vivo culture establishes that the cardiac twisting depends on intrinsic mechanisms and is independent from cardiac growth. Furthermore, genetic experiments indicate that looping requires proper tissue patterning. We conclude that cardiac looping involves twisting of the chambers around the atrioventricular canal, which requires correct tissue patterning by Tbx5a.

αß/γδ T cell lineage outcome is regulated by intrathymic cell localization and environmental signals.

αß and γδ T cells are two distinct sublineages that develop in the vertebrate thymus. Thus far, their differentiation from a common progenitor is mostly understood to be regulated by intrinsic mechanisms. However, the proportion of αß/γδ T cells varies in different vertebrate taxa. How this process is regulated in species that tend to produce a high frequency of γδ T cells is unstudied. Using an in vivo teleost model, the medaka, we report that progenitors first enter a thymic niche where their development into γδ T cells is favored. Translocation from this niche, mediated by chemokine receptor Ccr9b, is a prerequisite for their differentiation into αß T cells. On the other hand, the thymic niche also generates opposing gradients of the cytokine interleukin-7 and chemokine Ccl25a, and, together, they influence the lineage outcome. We propose a previously unknown mechanism that determines the proportion of αß/γδ lineages within species.

Retinal stem cells modulate proliferative parameters to coordinate post-embryonic morphogenesis in the eye of fish.

Combining clonal analysis with a computational agent based model, we investigate how tissue-specific stem cells for neural retina (NR) and retinal pigmented epithelium (RPE) of the teleost medaka (Oryzias latipes) coordinate their growth rates. NR cell division timing is less variable, consistent with an upstream role as growth inducer. RPE cells divide with greater variability, consistent with a downstream role responding to inductive signals. Strikingly, the arrangement of the retinal ciliary marginal zone niche results in a spatially biased random lineage loss, where stem- and progenitor cell domains emerge spontaneously. Further, our data indicate that NR cells orient division axes to regulate organ shape and retinal topology. We highlight an unappreciated mechanism for growth coordination, where one tissue integrates cues to synchronize growth of nearby tissues. This strategy may enable evolution to modulate cell proliferation parameters in one tissue to adapt whole-organ morphogenesis in a complex vertebrate organ.

Left/right asymmetric collective migration of parapineal cells is mediated by focal FGF signaling activity in leading cells.

The ability of cells to collectively interpret surrounding environmental signals underpins their capacity to coordinate their migration in various contexts, including embryonic development and cancer metastasis. One tractable model for studying collective migration is the parapineal, a left-sided group of neurons that arises from bilaterally positioned precursors that undergo a collective migration to the left side of the brain. In zebrafish, the migration of these cells requires Fgf8 and, in this study, we resolve how FGF signaling correlates with-and impacts the migratory dynamics of-the parapineal cell collective. The temporal and spatial dynamics of an FGF reporter transgene reveal that FGF signaling is activated in only few parapineal cells usually located at the leading edge of the parapineal during its migration. Overexpressing a constitutively active Fgf receptor compromises parapineal migration in wild-type embryos, while it partially restores both parapineal migration and mosaic expression of the FGF reporter transgene in fgf8-/- mutant embryos. Focal activation of FGF signaling in few parapineal cells is sufficient to promote the migration of the whole parapineal collective. Finally, we show that asymmetric Nodal signaling contributes to the restriction and leftwards bias of FGF pathway activation. Our data indicate that the first overt morphological asymmetry in the zebrafish brain is promoted by FGF pathway activation in cells that lead the collective migration of the parapineal to the left. This study shows that cell-state differences in FGF signaling in front versus rear cells is required to promote migration in a model of FGF-dependent collective migration.

Efficient single-copy HDR by 5' modified long dsDNA donors.

CRISPR/Cas9 efficiently induces targeted mutations via non-homologous-end-joining but for genome editing, precise, homology-directed repair (HDR) of endogenous DNA stretches is a prerequisite. To favor HDR, many approaches interfere with the repair machinery or manipulate Cas9 itself. Using Medaka we show that the modification of 5' ends of long dsDNA donors strongly enhances HDR, favors efficient single-copy integration by retaining a monomeric donor conformation thus facilitating successful gene replacement or tagging.

A 3D self-organizing multicellular epidermis model of barrier formation and hydration with realistic cell morphology based on EPISIM.

The epidermis and the stratum corneum (SC) as its outermost layer have evolved to protect the body from evaporative water loss to the environment. To morphologically represent the extremely flattened cells of the SC - and thereby the epidermal barrier - in a multicellular computational model, we developed a 3D biomechanical model (BM) based on ellipsoid cell shapes. We integrated the BM in the multicellular modelling and simulation platform EPISIM. We created a cell behavioural model (CBM) with EPISIM encompassing regulatory feedback loops between the epidermal barrier, water loss to the environment, and water and calcium flow within the tissue. This CBM allows a small number of stem cells to initiate self-organizing epidermal stratification, yielding the spontaneous emergence of water and calcium gradients comparable to experimental data. We find that the 3D in silico epidermis attains homeostasis most quickly at high ambient humidity, and once in homeostasis the epidermal barrier robustly buffers changes in humidity. Our model yields an in silico epidermis with a previously unattained realistic morphology, whose cell neighbour topology is validated with experimental data obtained from in vivo images. This work paves the way to computationally investigate how an impaired SC barrier precipitates disease.