RESUMEN

Orientational order, encoded in anisotropic fields, plays an important role during the development of an organism. A striking example of this is the freshwater polyp Hydra, where topological defects in the muscle fiber orientation have been shown to localize to key features of the body plan. This body plan is organized by morphogen concentration gradients, raising the question how muscle fiber orientation, morphogen gradients and body shape interact. Here, we introduce a minimal model that couples nematic orientational order to the gradient of a morphogen field. We show that on a planar surface, alignment to a radial concentration gradient can induce unbinding of topological defects, as observed during budding and tentacle formation in Hydra, and stabilize aster/vortex-like defects, as observed at a Hydra's mouth. On curved surfaces mimicking the morphologies of Hydra in various stages of development-from spheroid to adult-our model reproduces the experimentally observed reorganization of orientational order. Our results suggest how gradient alignment and curvature effects may work together to control orientational order during development and lay the foundations for future modeling efforts that will include the tissue mechanics that drive shape deformations.

Asunto(s)

RESUMEN

Morphogen gradients are a central concept in developmental biology. Their formation often involves the secretion of morphogens from a local source, that spread by diffusion in the cell field, where molecules eventually get degraded. This implies limits to both the time and length scales over which morphogen gradients can form which are set by diffusion coefficients and degradation rates. Towards the goal of identifying plausible mechanisms capable of extending the gradient range, we here use theory to explore properties of a cell-to-cell signaling relay. Inspired by the millimeter-scalewnt-expression and signaling gradients in flatworms, we consider morphogen-mediated morphogen production in the cell field. We show that such a relay can generate stable morphogen and signaling gradients that are oriented by a local, morphogen-independent source of morphogen at a boundary. This gradient formation can be related to an effective diffusion and an effective degradation that result from morphogen production due to signaling relay. If the secretion of morphogen produced in response to the relay is polarized, it further gives rise to an effective drift. We find that signaling relay can generate long-range gradients in relevant times without relying on extreme choices of diffusion coefficients or degradation rates, thus exceeding the limits set by physiological diffusion coefficients and degradation rates. A signaling relay is hence an attractive principle to conceptualize long-range gradient formation by slowly diffusing morphogens that are relevant for patterning in adult contexts such as regeneration and tissue turn-over.

Asunto(s)

RESUMEN

We propose a unified mechanism that reproduces the sequence of dynamical transitions observed during somitogenesis, the process of body segmentation during embryonic development, that is invariant across all vertebrate species. This is achieved by combining inter-cellular interactions mediated via receptor-ligand coupling with global spatial heterogeneity introduced through a morphogen gradient known to occur along the anteroposterior axis. Our model reproduces synchronized oscillations in the gene expression in cells at the anterior of the presomitic mesoderm as it grows by adding new cells at its posterior, followed by travelling waves and subsequent arrest of activity, with the eventual appearance of somite-like patterns. This framework integrates a boundary-organized pattern formation mechanism, which uses positional information provided by a morphogen gradient, with the coupling-mediated self-organized emergence of collective dynamics, to explain the processes that lead to segmentation.

Asunto(s)

RESUMEN

With the rise of new tools, from controlled genetic manipulations and optogenetics to improved microscopy, it is now possible to make clear, quantitative and reproducible measurements of biological processes. The humble fruit fly Drosophila melanogaster, with its ease of genetic manipulation combined with excellent imaging accessibility, has become a major model system for performing quantitative in vivo measurements. Such measurements are driving a new wave of interest from physicists and engineers, who are developing a range of testable dynamic models of active systems to understand fundamental biological processes. The reproducibility of the early Drosophila embryo has been crucial for understanding how biological systems are robust to unavoidable noise during development. Insights from quantitative in vivo experiments in the Drosophila embryo are having an impact on our understanding of critical biological processes, such as how cells make decisions and how complex tissue shape emerges. Here, to highlight the power of using Drosophila embryogenesis for quantitative biology, I focus on three main areas: (1) formation and robustness of morphogen gradients; (2) how gene regulatory networks ensure precise boundary formation; and (3) how mechanical interactions drive packing and tissue folding. I further discuss how such data has driven advances in modelling.

Asunto(s)

RESUMEN

In eukaryotic cells, mRNA molecules are coated with numerous RNA-binding proteins and so exist in ribonucleoproteins (mRNPs). The proteins associated with the mRNA regulate the fate of mRNA, including its localization, translation and decay. Before activation of translation, the mRNA does not display any template functions-it is masked. The coordinated activity of certain RNA-binding proteins determines the future fate of each mRNA individually. In embryo development, the temporal and spatial regulation of translation can cause a situation when the mRNA and the encoded protein are localized in different compartments and so the differentiation of the cells can be determined. The fundamentals of regulation of the mRNAs fate and functioning in nerves are similar to those already described for oo- and embryogenesis. Disorders in the mRNA masking and demasking result in the emergence of various diseases, in particular cancers and neuro-degenerative diseases.

Asunto(s)

RESUMEN

Development of multicellular organisms depends on the proper establishment of signaling information in space and time. Secreted molecules called morphogens form concentration gradients in space and provide positional information to differentiating cells within the organism. Although the key molecular components of morphogen pathways have been identified, how the architectures and key parameters of morphogen pathways control the properties of signaling gradients, such as their size, speed, and robustness to perturbations, remains challenging to study in developing embryos. Reconstituting morphogen gradients in cell culture provides an alternative approach to address this question. Here we describe the methodology for reconstituting Sonic Hedgehog (SHH) signaling gradients in mouse fibroblast cells. The protocol includes the design of morphogen sending and receiving cell lines, the setup of radial and linear gradients, the quantitative time-lapse imaging, and the data analysis. Similar approaches could potentially be applied to other cell-cell communication pathways.

Asunto(s)

RESUMEN

Over the past 5 years, several studies have begun to uncover the links between the classical signal transduction pathways and the physical mechanisms that are used to sculpt branched tissues. These advances have been made, in part, thanks to innovations in live imaging and reporter animals. With modern research tools, our conceptual models of branching morphogenesis are rapidly evolving, and the differences in branching mechanisms between each organ are becoming increasingly apparent. Here, we highlight four branched epithelia that develop at different spatial scales, within different surrounding tissues and via divergent physical mechanisms. Each of these organs has evolved to employ unique branching strategies to achieve a specialized final architecture.

Asunto(s)

RESUMEN

The notion that graded distributions of signals underlie the spatial organization of biological systems has long been a central pillar in the fields of cell and developmental biology. During morphogenesis, morphogens spread across tissues to guide development of the embryo. Similarly, a variety of dynamic gradients and pattern-forming networks have been discovered that shape subcellular organization. Here we discuss the principles of intracellular pattern formation by these intracellular morphogens and relate them to conceptually similar processes operating at the tissue scale. We will specifically review mechanisms for generating cellular asymmetry and consider how intracellular patterning networks are controlled and adapt to cellular geometry. Finally, we assess the general concept of intracellular gradients as a mechanism for positional control in light of current data, highlighting how the simple readout of fixed concentration thresholds fails to fully capture the complexity of spatial patterning processes occurring inside cells.

Asunto(s)

RESUMEN

Drosophila melanogaster embryos develop initially as a syncytium of totipotent nuclei and subsequently, once cellularized, undergo morphogenetic movements associated with gastrulation to generate the three somatic germ layers of the embryo: mesoderm, ectoderm, and endoderm. In this chapter, we focus on the first phase of gastrulation in Drosophila involving patterning of early embryos when cells differentiate their gene expression programs. This patterning process requires coordination of multiple developmental processes including genome reprogramming at the maternal-to-zygotic transition, combinatorial action of transcription factors to support distinct gene expression, and dynamic feedback between this genetic patterning by transcription factors and changes in cell morphology. We discuss the gene regulatory programs acting during patterning to specify the three germ layers, which involve the regulation of spatiotemporal gene expression coupled to physical tissue morphogenesis.

Asunto(s)

RESUMEN

Buffering variability in morphogen distribution is essential for reproducible patterning. A theoretically proposed class of mechanisms, termed "distal pinning," achieves robustness by combining local sensing of morphogen levels with global modulation of gradient spread. Here, we demonstrate a critical role for morphogen sensing by a gene enhancer, which ultimately determines the final global distribution of the morphogen and enables reproducible patterning. Specifically, we show that, while the pattern of Toll activation in the early Drosophila embryo is robust to gene dosage of its locally produced regulator, WntD, it is sensitive to a single-nucleotide change in the wntD enhancer. Thus, enhancer properties of locally produced WntD directly impinge on the global morphogen profile.

Asunto(s)

RESUMEN

Recently, it was shown that anterior-posterior patterning genes in the red flour beetle Tribolium castaneum are expressed sequentially in waves. However, in the fruit fly Drosophila melanogaster, an insect with a derived mode of embryogenesis compared to Tribolium, anterior-posterior patterning genes quickly and simultaneously arise as mature gene expression domains that, afterwards, undergo slight posterior-to-anterior shifts. This raises the question of how a fast and simultaneous mode of patterning, like that of Drosophila, could have evolved from a rather slow sequential mode of patterning, like that of Tribolium. In this paper, we propose a mechanism for this evolutionary transition based on a switch from a uniform to a gradient-mediated initialization of the gap gene cascade by maternal Hb. The model is supported by computational analyses and experiments.

Asunto(s)

RESUMEN

Multicellular animals and plants represent independent evolutionary experiments with complex multicellular bodyplans. Differences in their life history, a mobile versus sessile lifestyle, and predominant embryonic versus postembryonic development, have led to the evolution of highly different body plans. However, also many intriguing parallels exist. Extension of the vertebrate body axis and its segmentation into somites bears striking resemblance to plant root growth and the concomittant prepatterning of lateral root competent sites. Likewise, plant shoot phyllotaxis displays similarities with vertebrate limb and digit patterning. Additionally, both plants and animals use complex signalling systems combining systemic and local signals to fine tune and coordinate organ growth across their body. Identification of these striking examples of convergent evolution provides support for the existence of general design principles: the idea that for particular patterning demands, evolution is likely to arrive at highly similar developmental patterning mechanisms. Furthermore, focussing on these parallels may aid in identifying core mechanistic principles, often obscured by the highly complex nature of multiscale patterning processes.

Asunto(s)

RESUMEN

Embryonic development is heavily dependent on temporally and spatially restricted gene expression. Spatially resolved measurements of gene expression are therefore crucial for identifying novel regulators and the understanding of their function. However, in situ methods do not resolve global gene expression, and sequencing-based methods usually do not provide spatial information. Here, we describe tomo-seq, a method that combines classical histological sectioning of embryos or tissues with a highly sensitive RNA-sequencing technique. Application of tomo-seq to zebrafish embryos allows reconstructing the spatial gene expression of thousands of genes.

Asunto(s)

RESUMEN

Soluble signal gradients play an important role in organ patterning, cell migration, and differentiation. Currently, signal gradients in 2D cell culture are realized using microfluidics and here cells are exposed to high and nonphysiological shear stress. Tissue morphogenesis (organogenesis) however occurs in 3D and therefore there is a need for simple and practical systems to impose gradients to cells dispersed in 3D matrix. Herein, a 3D gradient generator based on passive diffusion elements that recapitulates interstitial flow and is capable of imposing predictable gradients over long length scales (6 mm) lasting up to 48 h to cells dispersed in a hydrogel environment is reported. Using recombinant human WNT3A (rhWNT3A), the spatiotemporal activation of the canonical WNT pathway in human epithelial kidney cells and human mesenchymal stems cells expressing a green fluorescence protein reporter on a transcription factor/lymphoid enhancer-binding factor (TCF/LEF) promoter is demonstrated. By refining computation models based on experimental findings, the diffusion coefficient of rhWNT3A in presence of human cells in 3D is determined. Furthermore, the formation of rhBMP4 gradients is visualized using immunohistochemistry by staining for phospho-SMAD1/5, the downstream targets of the bone morphogenetic protein (BMP) pathway. The simplicity of the gradient generator is expected to spur its adoption in studying developmental biology paradigms in vitro.

RESUMEN

Morphogens are proteins that form concentration gradients in embryos and developing tissues, where they act as postal codes, providing cells with positional information and allowing them to behave accordingly. Bicoid was the first discovered morphogen, and remains one of the most studied. It regulates segmentation in flies, forming a striking exponential gradient along the anterior-posterior axis of early Drosophila embryos, and activating the transcription of multiple target genes in a concentration-dependent manner. In this review, the work done by us and by others to characterize the mobility of Bicoid in D. melanogaster embryos is presented. The central role played by the diffusion of Bicoid in both the establishment of the gradient and the activation of target genes is discussed, and placed in the context of the need for these processes to be all at once rapid, precise and robust. The Bicoid system, and morphogen gradients in general, remain amongst the most amazing examples of the coexistence, often observed in living systems, of small-scale disorder and large-scale spatial order. This article is part of a Special Issue entitled: Biophysics in Canada, edited by Lewis Kay, John Baenziger, Albert Berghuis and Peter Tieleman.

Asunto(s)

RESUMEN

Morphogen gradients determine tissue pattern by triggering differential cell responses to distinct morphogen concentrations. The strict quantitative dependence of the emerging patterns on morphogen distribution raises the challenge of buffering variability in morphogen profile to ensure a reproducible outcome. We describe the underlying principles of two modules for buffering morphogen distribution: buffering morphogen amplitude by storing excess morphogen in a limited spatial region, and buffering morphogen spread by pinning morphogen levels at a distal position through global feedback that adjusts morphogen diffusion or degradation across the tissue. We also present concrete examples of patterning systems that implement these modules.

Asunto(s)

RESUMEN

Three-dimensional organoids derived from human pluripotent stem cell (hPSC) derivatives have become widely used in vitro models for studying development and disease. Their ability to recapitulate facets of normal human development during in vitro morphogenesis produces tissue structures with unprecedented biomimicry. Current organoid derivation protocols primarily rely on spontaneous morphogenesis processes to occur within 3-D spherical cell aggregates with minimal to no exogenous control. This yields organoids containing microscale regions of biomimetic tissues, but at the macroscale (i.e. 100's of microns to millimeters), the organoids' morphology, cytoarchitecture, and cellular composition are non-biomimetic and variable. The current lack of control over in vitro organoid morphogenesis at the microscale induces aberrations at the macroscale, which impedes realization of the technology's potential to reproducibly form anatomically correct human tissue units that could serve as optimal human in vitro models and even transplants. Here, we review tissue engineering methodologies that could be used to develop powerful approaches for instructing multiscale, 3-D human organoid morphogenesis. Such technological mergers are critically needed to harness organoid morphogenesis as a tool for engineering functional human tissues with biomimetic anatomy and physiology. STATEMENT OF SIGNIFICANCE: Human PSC-derived 3-D organoids are revolutionizing the biomedical sciences. They enable the study of development and disease within patient-specific genetic backgrounds and unprecedented biomimetic tissue microenvironments. However, their uncontrolled, spontaneous morphogenesis at the microscale yields inconsistences in macroscale organoid morphology, cytoarchitecture, and cellular composition that limits their standardization and application. Integration of tissue engineering methods with organoid derivation protocols could allow us to harness their potential by instructing standardized in vitro morphogenesis to generate organoids with biomimicry at all scales. Such advancements would enable the use of organoids as a basis for 'next-generation' tissue engineering of functional, anatomically mimetic human tissues and potentially novel organ transplants. Here, we discuss critical aspects of organoid morphogenesis where application of innovative tissue engineering methodologies would yield significant advancement towards this goal.

Asunto(s)

RESUMEN

How the shape and size of tissues and organs is regulated during development is a major question in developmental biology. Such regulation relies upon both intrinsic cues (such as signaling networks) and extrinsic inputs (such as from neighboring tissues). Here, we focus on pattern formation and organ development during Drosophila embryogenesis. In particular, we outline the importance of both biochemical and mechanical tissue-tissue interactions in size regulation. We describe how the Drosophila embryo can potentially provide novel insights into how shape and size are regulated during development. We focus on gene expression boundary scaling in the early embryo and how size is regulated in three organs (hindgut, trachea, and ventral nerve cord) later in development, with particular focus on the role of tissue-tissue interactions. Overall, we demonstrate that Drosophila embryogenesis provides a suitable model system for studying spatial and temporal scaling and size control in vivo.

Asunto(s)

RESUMEN

Robust development of biological organisms in the presence of genetic and epi-genetic perturbations is important for time spans short relative to evolutionary time. Gradients of receptor bound signaling morphogens are responsible for patterning formation and development. A variety of inhibitors for reducing ectopic signaling activities are known to exist and their specific role in down-regulating the undesirable ectopic activities reasonably well understood. However, how a developing organism manages to adjust inhibition/stimulation in response to genetic and/or environmental changes remains to be uncovered. The need to adjust for ectopic signaling activities requires the presence of one or more feedback mechanisms to stimulate the needed adjustment. As the ultimate effect of many inhibitors (including those of the nonreceptor type) is to reduce the availability of signaling morphogens for binding with signaling receptors, a negative feedback on signaling morphogen synthesis rate based on a root-mean-square measure of the spatial distribution of signaling concentration offers a simple approach to robusness and has been demonstrated to be effective in a proof-of-concept implementation. In this paper, we complement the previous investigation of feedback in steady state by examining the effect of one or more feedback adjustments during the transient phase of the biological development.

RESUMEN

It is long established that the graded distribution of Dorsal transcription factor influences spatial domains of gene expression along the dorsoventral (DV) axis of Drosophila melanogaster embryos. However, the more recent realization that Dorsal levels also change with time raises the question of whether these dynamics are instructive. An overview of DV axis patterning is provided, focusing on new insights identified through quantitative analysis of temporal changes in Dorsal target gene expression from one nuclear cycle to the next ('steps'). Possible roles for the stepwise progression of this patterning program are discussed including (i) tight temporal regulation of signaling pathway activation, (ii) control of gene expression cohorts, and (iii) ensuring the irreversibility of the patterning and cell fate specification process.

Asunto(s)