The zebrafish presomitic mesoderm elongates through compression-extension

In vertebrate embryos the presomitic mesoderm become progressively segmented into somites at the anterior end while extending along the anterior-posterior axis. A commonly adopted model to explain how this tissue elongates is that of posterior growth, driven in part by the addition of new cells from uncommitted progenitor populations in the tailbud. However, in zebrafish, much of somitogenesis is associated with an absence of overall volume increase and posterior progenitors do not contribute new cells until the final stages of somitogenesis. Here, we perform a comprehensive 3D morphometric analysis of the paraxial mesoderm and reveal that extension is linked to a volumetric decrease, compression in both dorsal-ventral and medio-lateral axes, and an increase in cell density. We also find that individual cells decrease in their cell volume over successive somite stages. Live cell tracking confirms that much of this tissue deformation occurs within the presomitic mesoderm progenitor zone and is associated with non-directional rearrangement. Furthermore, unlike the trunk somites that are laid down during gastrulation, tail somites develop from a tissue that can continue to elongate in the absence of functional PCP signalling. Taken together, we propose a compression-extension mechanism of tissue elongation that highlights the need to better understand the role of tissue intrinsic and extrinsic forces play in regulating morphogenesis.

Morphogenetic coupling leads to pattern emergence in the presomitic mesoderm

Pattern formation in development has been principally studied in tissues that are not undergoing extensive cellular rearrangement. However, in most developmental contexts, gene expression domains emerge as cells re-arrange their spatial positions within the tissue, providing an additional, and seldom explored, level of complexity to the process of pattern formation in vivo. To investigate this issue, we addressed the regulation of TBox expression in the presomitic mesoderm (PSM) as this tissue develops in zebrafish embryos. Here, cells must differentiate in a manner that leads to well-defined spatial gene expression domains along the tissue while undergoing rapid movements to generate axial length. We find that in vivo, mesoderm progenitors undergo TBox differentiation over a broad range of time scales while in vitro their differentiation is simultaneous. By reverse-engineering a gene regulatory network (GRN) to recapitulate TBox gene expression, we were able to predict the population-level differentiation dynamics observed in culture, but not in vivo. In order to address this discrepancy in differentiation dynamics we developed a ‘Live Modelling’ framework that allowed us to simulate the GRN on 3D tracking data generated from large-scale time-lapse imaging datasets of the developing PSM. Once the network was simulated on a realistic representation of the cells’ morphogenetic context, the model was able to recapitulate the range of differentiation time scales observed in vivo, and revealed that these were necessary for TBox gene expression patterns to emerge correctly at the level of the tissue. This work thus highlights a previously unappreciated role for cell movement as a driver of pattern formation in development.

Generation of a new Tbx6-inducible reporter mouse line to trace presomitic mesoderm derivatives throughout development and in adults

ABSTRACTThe presomitic mesoderm (PSM) is initially an unsegmented structure localized on each side of the neural tube of the developing embryo, which progressively segments to form the somites. The somites will segregate and partition to generate the dorsal dermomyotome and the ventral sclerotome. Endothelial and myogenic cells of both the trunk and limbs are derived from the somites. There is a lack of efficient reporter mouse models to label and trace the PSM derivatives, despite their crucial contribution to many developmental processes. In this study, we generated a tamoxifen inducible transgenic Tbx6 mouse line, Tg(Tbx6_Cre/ERT2)/ROSA-eYFP, to tag and follow PSM-derivatives from early embryonic stages until adulthood. After induction, endothelial and myogenic cells can be easily identified within the trunk and limbs with proper expression patterns. Since our Tg(Tbx6_Cre/ERT2)/ROSA-eYFP model allows to permanently label the PSM-derived cells, their progeny can be studied at long-term, opening the possibility to perform lineage tracing of stem cells upon aging.

Cellular Synchronisation through Unidirectional and Phase-Gated Signalling

Multiple natural and artificial oscillator systems achieve synchronisation when oscillators are coupled. The coupling mechanism, essentially the communication between oscillators, is often assumed to be continuous and bidirectional. However, the cells of the presomitic mesoderm synchronise their gene expression oscillations through Notch signalling, which is intermittent and directed from a ligand-presenting to a receptor-presenting cell. Motivated by this mode of communication we present a phase-gated and unidirectional coupling mechanism. We identify conditions under which it can successfully bring two or more oscillators to cycle in-phase. In the presomitic mesoderm we observed the oscillatory dynamics of two synchronizing cell populations and record one population halting its pace while the other keeps undisturbed, as would be predicted from our model. For the same system another important prediction, convergence to a specific range of phases upon synchronisation is also confirmed. Thus, the proposed mechanism accurately describes the coordinated oscillations of the presomitic mesoderm cells and provides an alternative framework for deciphering synchronisation.

In vivo characterization of chick embryo mesoderm by optical coherence tomography assisted microindentation

Embryos are growing organisms with highly heterogeneous properties in space and time. Understanding the mechanical properties is a crucial prerequisite for the investigation of morphogenesis. During the last ten years, new techniques have been developed to evaluate the mechanical properties of biological tissues in vivo. To address this need, we employed a new instrument that, via the combination of micro-indentation with Optical Coherence Tomography (OCT), allows us to determine both, the spatial distribution of mechanical properties of chick embryos and the structural changes in real-time provided by OCT. We report here the stiffness measurements on live chicken mesoderm during somite formation, from the mesenchymal tailbud to the epithelialized somites. The storage modulus of the mesoderm increases from (176±18) Pa in the tail up to (716±117) Pa in the somitic region. The midline has a storage modulus of (947±111) Pa in the caudal presomitic mesoderm, indicating a stiff rod along the body axis, which thereby mechanically supports the surrounding tissue. The difference in stiffness between midline and presomitic mesoderm decreases as the mesoderm forms somites. The viscoelastic response of the somites develops further until somite IV, which is commensurate with the slow process of epithelization of somites between S0 and SIV.Overall, this study provides an efficient method for the biomechanical characterization of soft biological tissues in vivo and shows that the mechanical properties strongly relate to different morphological features of the investigated regions.

Intrinsic noise, Delta-Notch signalling and delayed reactions promote sustained, coherent, synchronized oscillations in the presomitic mesoderm

Using a stochastic individual-based modelling approach, we examine the role that Delta-Notch signalling plays in the regulation of a robust and reliable somite segmentation clock. We find that not only can Delta-Notch signalling synchronize noisy cycles of gene expression in adjacent cells in the presomitic mesoderm (as is known), but it can also amplify and increase the coherence of these cycles. We examine some of the shortcomings of deterministic approaches to modelling these cycles and demonstrate how intrinsic noise can play an active role in promoting sustained oscillations, giving rise to noise-induced quasi-cycles. Finally, we explore how translational/transcriptional delays can result in the cycles in neighbouring cells oscillating in anti-phase and we study how this effect relates to the propagation of noise-induced stochastic waves.