Dynamic Expression oflunatic fringeSuggests a Link betweennotchSignaling and an Autonomous Cellular Oscillator Driving Somite Segmentation

AbstractThe metameric organization of the vertebrate trunk is a characteristic feature of all members of this phylum. The origin of this metamerism can be traced to the division of paraxial mesoderm into individual units, termed somites, during embryonic development. Despite the identification of somites as the first overt sign of segmentation in vertebrates well over 100 years ago, the mechanism(s) underlying somite formation remain poorly understood. Recently, however, several genes have been identified which play prominent roles in orchestrating segmentation, including the novel secreted factorlunatic fringe.To gain further insight into the mechanism by whichlunatic fringecontrols somite development, we have conducted a thorough analysis oflunatic fringeexpression in the unsegmented paraxial mesoderm of chick embryos. Here we report thatlunatic fringeis expressed predominantly in somite −II, where somite I corresponds to the most recently formed somite and somite −I corresponds to the group of cells which will form the next somite. In addition, we show thatlunatic fringeis expressed in a highly dynamic manner in the chick segmental plate prior to somite formation and thatlunatic fringeexpression cycles autonomously with a periodicity of somite formation. Moreover, the murine ortholog oflunatic fringeundergoes a similar cycling expression pattern in the presomitic mesoderm of somite stage mouse embryos. The demonstration of a dynamic periodic expression pattern suggests thatlunatic fringemay function to integrate notch signaling to a cellular oscillator controlling somite segmentation

A propos de gradients et d'oscillations: le rôle de la voie de signalisation Wnt dans la formation des somites au cours du développement embryonnaire

A fundamental characteristic of all vertebrates is the metameric organization of their body plan, best exemplified by the vertebral column. This organization originates during embryonic development and the emergence of the vertebrae precursor, the somites, which form from the paraxial presomitic mesoderm (PSM). The discovery of oscillatory transcriptional activity of the Notch-signaling pathway within the PSM supports the existence of an embryological clock, the segmentation clock, which is responsible in controlling the periodic formation of somites. Here, I present the discovery of oscillatory mRNA expression of Axin2, a target of the Wnt-signaling pathway, in the PSM. This establishes a novel link between Wnt signaling and the segmentation clock. I then describe the generation of a two-photon, real-time imaging technology that allows visualizing segmentation clock activity in realtime in living mouse embryos. This enables us to directly measure oscillation parameters in the PSM. Finally, I describe the discovery of a -catenin protein gradient in the PSM. Using conditional gene targeting experiments, we establish that this gradient controls PSM maturation and differentiation. Moreover, this gradient represents a permissive signal that allows segmentation clock activity to occur. Based on the results presented, I propose a novel model for somite formation.Une caractéristique fondamentale des vertébrés est leur organisation métamerique, visible au niveau de la colonne vertébrale. Cette organisation se met en place au cours du développement embryonnaire et l'émergence des précurseurs des vertèbres, les somites, formés à partir du mésoderm paraxial présomitique (PSM). Depuis la découverte d'une activité transcriptionelle oscillatoire de la voie de signalisation Notch dans le PSM, on propose que cette activité oscillatoire représente l'action d'une horloge embryonnaire, l'horloge de segmentation, responsable de contrôler la formation des somites de façon périodique. Dans cette étude, je présente la découverte d'une nouvelle association de la voie Wnt avec l'horloge de segmentation en décrivant l'activité oscillatoire de Axin2, une cible de la voie Wnt, dans le PSM. Ensuite, j'ai mise en place un system d'imagerie biphoton qui nous permet d'observer l'action de l'horloge de segmentation en temps réel et in vivo dans les embryons de souris. Ce système permet de mesurer directement les paramètres des oscillations. Ultérieurement je décris la découverte d'un gradient d'expression de la protéine -caténine dans le PSM. A l'aide d'expériences de recombinaisons homologues conditionnelles nous avons déterminé que ce gradient de - caténine contrôle la maturation et différentiation des cellules dans le PSM. De plus, ce gradient constitue un signal essentiel et permissif pour les oscillations de l'horloge de segmentation. En conclusion, je propose un nouveau modèle de mise en place des somites incorporant les résultats présentés

About gradients and oscillations (the role of the Wnt-signaling pathway in somite formation during embryonic development)

PARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Metabolic control of cellular differentiation

The role of metabolic rewiring during cellular differentiation is under intense investigation. Reporting recently in Science, Peng et al. (2016) found that activation of glycolysis supports T helper cell differentiation by controlling acetyl-coA and histone acetylation levels, identifying a link between metabolic state and epigenetic control of gene activity

Signaling Gradients during Paraxial Mesoderm Development

The sequential formation of somites along the anterior-posterior axis is under control of multiple signaling gradients involving the Wnt, FGF, and retinoic acid (RA) pathways. These pathways show graded distribution of signaling activity within the paraxial mesoderm of vertebrate embryos. Although Wnt and FGF signaling show highest activity in the posterior, unsegmented paraxial mesoderm (presomitic mesoderm [PSM]), RA signaling establishes a countergradient with the highest activity in the somites. The generation of these graded activities relies both on classical source-sink mechanisms (for RA signaling) and on an RNA decay mechanism (for FGF signaling). Numerous studies reveal the tight interconnection among Wnt, FGF, and RA signaling in controlling paraxial mesoderm differentiation and in defining the somite-forming unit. In particular, the relationship to a molecular oscillator acting in somite precursors in the PSM—called the segmentation clock—has been recently addressed. These studies indicate that high levels of Wnt and FGF signaling are required for the segmentation clock activity. Furthermore, we discuss how these signaling gradients act in a dose-dependent manner in the progenitors of the paraxial mesoderm, partly by regulating cell movements during gastrulation. Finally, links between the process of axial specification of vertebral segments and Hox gene expression are discussed

Wnt3a plays a major role in the segmentation clock controlling somitogenesis

The vertebral column derives from somites generated by segmentation of presomitic mesoderm (PSM). Somitogenesis involves a molecular oscillator, the segmentation clock, controlling periodic Notch signaling in the PSM. Here, we establish a novel link between Wnt/β-catenin signaling and the segmentation clock. Axin2, a negative regulator of the Wnt pathway, is directly controlled by Wnt/β-catenin and shows oscillating expression in the PSM, even when Notch signaling is impaired, alternating with Lfng expression. Moreover, Wnt3a is required for oscillating Notch signaling activity in the PSM. We propose that the segmentation clock is established by Wnt/β-catenin signaling via a negative-feedback mechanism and that Wnt3a controls the segmentation process in vertebrates

WNT signaling, in synergy with T/TBX6, controls Notch signaling by regulating Dll1 expression in the presomitic mesoderm of mouse embryos

Notch signaling in the presomitic mesoderm (psm) is critical for somite formation and patterning. Here, we show that WNT signals regulate transcription of the Notch ligand Dll1 in the tailbud and psm. LEF/TCF factors cooperate with TBX6 to activate transcription from the Dll1 promoter in vitro. Mutating either T or LEF/TCF sites in the Dll1 promoter abolishes reporter gene expression in vitro as well as in the tail bud and psm of transgenic embryos. Our results indicate that WNT activity, in synergy with TBX6, regulates Dll1 transcription and thereby controls Notch activity, somite formation, and patterning

Spatiotemporal analysis of a glycolytic activity gradient linked to mouse embryo mesoderm development

How metabolism is rewired during embryonic development is still largely unknown, as it remains a major technical challenge to resolve metabolic activities or metabolite levels with spatiotemporal resolution. Here, we investigated metabolic changes during development of organogenesis-stage mouse embryos, focusing on the presomitic mesoderm (PSM). We measured glycolytic labeling kinetics from 13C-glucose tracing experiments and detected elevated glycolysis in the posterior, more undifferentiated PSM. We found evidence that the spatial metabolic differences are functionally relevant during PSM development. To enable real-time quantification of a glycolytic metabolite with spatiotemporal resolution, we generated a pyruvate FRET-sensor reporter mouse line. We revealed dynamic changes in cytosolic pyruvate levels as cells transit toward a more anterior PSM state. Combined, our approach identifies a gradient of glycolytic activity across the PSM, and we provide evidence that these spatiotemporal metabolic changes are intrinsically linked to PSM development and differentiation