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During vertebrate embryogenesis, the rhythmic and sequential segmentation of the body axis is regulated by an oscillating genetic network termed the segmentation clock. We describe a new dynamic model for the core pace-making circuit of the zebrafish segmentation clock based on a systematic biochemical investigation of the network's topology and precise measurements of somitogenesis dynamics in novel genetic mutants. We show that the core pace-making circuit consists of two distinct negative feedback loops, one with Her1 homodimers and the other with Her7:Hes6 heterodimers, operating in parallel. To explain the observed single and double mutant phenotypes of her1, her7, and hes6 mutant embryos in our dynamic model, we postulate that the availability and effective stability of the dimers with DNA binding activity is controlled in a "dimer cloud" that contains all possible dimeric combinations between the three factors. This feature of our model predicts that Hes6 protein levels should oscillate despite constant hes6 mRNA production, which we confirm experimentally using novel Hes6 antibodies. The control of the circuit's dynamics by a population of dimers with and without DNA binding activity is a new principle for the segmentation clock and may be relevant to other biological clocks and transcriptional regulatory networks. © 2012 Schröter et al. Fil: Schroter, Christian. University of Cambridge; Estados Unidos Fil: Ares, Saúl. Max Planck Institute For The Physics Of Complex Systems; Alemania Fil: Morelli, Luis Guillermo. Universidad de Buenos Aires; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Física de Buenos Aires. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Física de Buenos Aires; Argentina Fil: Isakova, Alina. Ecole Polytechnique Federale de Lausanne; Francia Fil: Hens, Korneel. Ecole Polytechnique Federale de Lausanne; Francia Fil: Soroldoni, Daniele. Max Planck Institute Of Molecular Cell Biology And Genetics; Alemania Fil: Gajewski, Martin. Universitat zu Köln; Alemania Fil: Jülicher, Frank. Max Planck Institute For The Physics Of Complex Systems; Alemania Fil: Maerkl, Sebastian J.. Ecole Polytechnique Federale de Lausanne; Francia Fil: Deplancke, Bart. Ecole Polytechnique Federale de Lausanne; Francia Fil: Oates, Andrew C.. Max Planck Institute Of Molecular Cell Biology And Genetics; Alemania

The segmentation of the vertebrate body plan during embryonic development is a rhythmic and sequential process governed by genetic oscillations. These genetic oscillations give rise to traveling waves of gene expression in the segmenting tissue. Here we present a minimal continuum theory of vertebrate segmentation that captures the key principles governing the dynamic patterns of gene expression including the effects of shortening of the oscillating tissue. We show that our theory can quantitatively account for the key features of segmentation observed in zebrafish, in particular the shape of the wave patterns, the period of segmentation and the segment length as a function of time. Fil: Jörg, David J. Max Planck Institute for the Physics of Complex Systems; Alemania Fil: Morelli, Luis Guillermo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Física de Buenos Aires. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Física de Buenos Aires; Argentina Fil: Soroldoni, Daniele. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Física de Buenos Aires. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Física de Buenos Aires; Argentina Fil: Oates, Andrew C.. Francis Crick Institute, Mill Hill Laboratory; Alemania Fil: Jülicher, Frank. Max Planck Institute for the Physics of Complex Systems; Alemania

The ability to quantify transcriptional dynamics in individual cells via live imaging has revolutionized our understanding of gene regulation. However, such measurements are lacking in the context of vertebrate embryos. We addressed this deficit by applying MS2-MCP mRNA labeling to the quantification of transcription in zebrafish, a model vertebrate. We developed a platform of transgenic organisms, light sheet fluorescence microscopy, and optimized image analysis that enables visualization and quantification of MS2 reporters. We used these tools to obtain the first single-cell, real-time measurements of transcriptional dynamics of the segmentation clock. Our measurements challenge the traditional view of smooth clock oscillations and instead suggest a model of discrete transcriptional bursts that are organized in space and time. Together, these results highlight how measuring single-cell transcriptional activity can reveal unexpected features of gene regulation and how this data can fuel the dialogue between theory and experiment.