Development Growth & Differentiation最新文献

Pleurodeles waltl is coming to light as a model animal, especially in regeneration studies, but deep studies on the molecular mechanisms have been limited due to the absence of primary tissue cells for wide usage. Therefore, we aimed to grow primary cells from limb tissue of P. waltl for in vitro experiments. Limb tissues were cut into small pieces and seeded as “explants” on culture dishes coated with fibronectin and gelatin. Compared to the control without coating, both fibronectin and gelatin supported quicker outgrowth of cells from explants and faster cell adhesion, and fibronectin showed significantly better performance than gelatin. Interestingly, the doubling time of cells on fibronectin- and gelatin-coated surfaces was almost the same (42.39 ± 2.79 h vs. 42.91 ± 3.69 h) and was not significantly different from that on non-coated plates (49.64 ± 3.63 h). The cryopreserved cells were successfully recovered and showed a multiplication capacity that was similar to that of fresh cells. Senescent cells were barely detected even after long-term sub-culture (>15 passages). Moreover, enhanced fluorescence of MitoSOX™ Red in cells under H₂O₂ exposure confirmed the respondence to chemical stimuli. Collectively, our results show that we are able to grow enough good-quality cells from P. waltl limb tissue for in vitro experiments, and fibronectin coating provides the best biocompatible environment for cell outgrowth and attachment.

We review several mathematical models and concepts in developmental biology that have been established over the last decade. (1) Feedback vertex set: Ascidian embryos contain cells of seven types, and cell fate is controlled by ~100 interacting genes. The “feedback vertex set” of the directed graph of the gene regulatory network consists of a small number of genes. By experimentally manipulating them, we can differentiate cells into any cell type. (2) Tissue deformation: Describing morphological changes in tissues and relating them to gene expression and other cellular processes is key in understanding morphogenesis. Expansion and anisotropy of the tissue are described by a “deformation tensor” at each location. A study on chick limb bud formation revealed that both the volume growth rate and anisotropy in deformation differed significantly between locations and stages. (3) Mechanobiology: Forces operating on each cell may alter cell shape and gene expression, which may subsequently exert forces on their surroundings. Measurements of force, tissue shape, and gene expression help us understand autonomous tissue deformation. (4) Adaptive design of development: An optimal growth schedule in fluctuating environments explains the growth response to starvation in Drosophila larvae. Adaptive placement of morphogen sources makes development robust to noises.

Cell fate decisions emerge as a consequence of a complex set of gene regulatory networks. Models of these networks are known to have more parameters than data can determine. Recent work, inspired by Waddington's metaphor of a landscape, has instead tried to understand the geometry of gene regulatory networks. Here, we describe recent results on the appropriate mathematical framework for constructing these landscapes. This allows the construction of minimally parameterized models consistent with cell behavior. We review existing examples where geometrical models have been used to fit experimental data on cell fate and describe how spatial interactions between cells can be understood geometrically.

Although the heart is one of the most important organs for animal survival, its regenerative capacity varies among animal species. Notably, adult mammals cannot regenerate their hearts after damage such as acute myocardial infarction. In contrast, some vertebrate animals can regenerate the heart throughout their lives. Cross-species comparative studies are important to understand the full picture of cardiac regeneration in vertebrates. Among the animal species able to regenerate the heart, some urodele amphibians, such as newts, possess a remarkable capacity for this process. Standardized methods of inducing cardiac regeneration in the newt are needed as a platform for studies comparing newts and other animal models. The procedures presented here describe amputation and cryo-injury techniques for the induction of cardiac regeneration in Pleurodeles waltl, an emerging model newt species. Both procedures consist of simplified steps that require no special equipment. We additionally show some examples of the regenerative process obtained using these procedures. This protocol has been developed for P. waltl. However, these methods are also expected to be applicable to other newt and salamander species, facilitating comparative research with other model animals.

Mechanosensing, the active responses of cells to the mechanics on multiple scales, plays an indispensable role in regulating cell behaviors and determining the fate of biological entities such as tissues and organs. Here, I aim to give a pedagogical illustration of the fundamental concepts of soft matter physics that aid in understanding biomechanical phenomena from the scale of tissues to proteins. Examples of up-to-date research are introduced to elaborate these concepts. Challenges in applying physics models to biology have also been discussed for biologists and physicists to meet in the field of mechanobiology.

Cryptochromes (CRYs) are multifunctional molecules that act as a circadian clock oscillating factor, a blue-light sensor, and a light-driven magnetoreceptor. Cry genes are classified into several groups based on the evolutionary relationships. Cryptochrome 6 gene (Cry6) is present in invertebrates and lower vertebrates such as amphibians and fishes. Here we identified a Cry6 ortholog in Xenopus tropicalis (XtCry6). XtCRY6 retains a conserved long N-terminal extension (termed CRY N-terminal extension; CNE) that is not found in any CRY in the other groups. A structural prediction suggested that CNE contained unique structures; a tetrahelical fold structure topologically related to KaiA/RbsU domain, overlapping nuclear- and nucleolar-localizing signals (NLS/NoLS), and a novel motif (termed DI-UIM) overlapping a double-sided ubiquitin-interacting motif (DUIM) and an inverted ubiquitin-interacting motif (IUIM). Potential activities of the NLS/NoLS and DI-UIM were examined to infer the molecular function of XtCRY6. GFP-NLS/NoLS fusion protein exogenously expressed in HEK293 cells was mostly observed in the nucleolus, while GFP-XtCRY6 was observed in the cytoplasm. A glutathione S-transferase (GST) pull-down assay suggested that the DI-UIM physically interacts with polyubiquitin. Consistently, protein docking simulations implied that XtCRY6 DI-UIM binds two ubiquitin molecules in a relationship of a twofold rotational symmetry with the symmetry axis parallel or perpendicular to the DI-UIM helix. These results strongly suggested that XtCRY6 does not function as a circadian transcriptional repressor and that it might have another function such as photoreceptive molecule regulating light-dependent protein degradation or gene expression through a CNE-mediated interaction with ubiquitinated proteins in the cytoplasm and/or nucleolus.

The mammalian secondary palate develops through complex processes including palatal shelf growth, elevation, and fusion. Palatal shelf elevation is a process accompanied by large-scale morphological changes over a short period. The elevation pattern changes along the anterior–posterior axis; the anterior region elevates by the “flip-up” model, and the middle and posterior regions reorient through the “flow” model. However, the mechanisms of both models are unclear because of the rapid progression of the elevation in utero. To observe palatal elevation in real time in detail, we aimed to establish a live imaging method using explants of the anterior region of the palatal shelf in mouse embryos before the beginning of elevation. Changes in the degree of shelf orientation were measured, which showed that the palatal shelf continuously changed shape toward the lingual side. The changes in the angle between the lingual and buccal bases of the palatal shelf were different; the morphological change at the lingual side resulted in a more acute angle, and the change at the buccal side resulted in a more obtuse angle. The morphological changes of the lingual and buccal sides occurred nearly simultaneously, suggesting that the anterior region of the palatal shelf in vitro elevated according to the “flip-up” model. This live imaging method enables the continuous observation of palatal shelf elevation and provides new insights into palatogenesis.

This is a meeting report of “The workshop of research and techniques using next-generation sequencing (NGS) for developmental biology 2022.” About 20 researchers attended the workshop. We discuss their NGS studies and techniques.

The Third Franco-Japanese Developmental Biology Meeting has been held to reinforce the partnership and facilitate the international collaborations and the academic exchanges between the French Society of Developmental Biology (SFBD) and the Japanese Society of Developmental Biology (JSDB). Nearly 200 participants attended, and we actively discussed interdisciplinary knowledge throughout the meeting period. The meeting realized not only the tightest connection between these two communities ever, but also a great contribution to international interdisciplinary academic communications and scientific communications with the public. In order to celebrate and maintain this massive momentum to push forward the entire field of developmental biology, we share the atmosphere with the entire international community and next generations through this report.