Research Article

Contact area–dependent cell communication and the morphological invariance of ascidian embryogenesis

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Science  10 Jul 2020:
Vol. 369, Issue 6500, eaar5663
DOI: 10.1126/science.aar5663

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Cell-cell contacts specify cell fate

Ascidians, or sea squirts, are marine invertebrate filter feeders with highly reproducible cellular events and invariant embryonic cell lineages. Guignard et al. studied the ascidian embryo to address the determinants of this cellular reproducibility. They introduce computational methods for the robust and automated segmentation, tracking, and analysis of whole-cell behaviors in high-throughput light-sheet microscopy datasets. This work shows that cell induction can be controlled by the contact area among cells. The range of cell signaling is proposed to set the scale at which animal embryonic reproducibility is observed. A high level of reproducibility of embryonic geometries may also counter-intuitively lift constraints on genome evolution, thereby contributing to the rapid molecular evolution observed in ascidians.

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Structured Abstract

INTRODUCTION

Within each animal species, embryonic development is highly reproducible, ensuring the faithful production of a complex organism with precisely arranged and shaped organs. In most animal embryos, reproducibility is found at the tissue scale, the behaviors of individual cells being stochastic beyond the first cell divisions. Ascidians, a group of marine invertebrate chordates, show an extreme form of embryoni­­c reproducibility: Homologous cells can be found across individual embryos, and early emb­­ryonic cell lineages are considered invariant. Embryonic geometries are even conserved between species, which diverged 400 million years ago and have very dissimilar genomes. Because of their evolutionary conservation of early embryonic development and ability to buffer genetic divergence, ascidians constitute attractive model systems to study the mechanisms driving cellular reproducibility.

RATIONALE

To quantify embryonic reproducibility in the ascidian Phallusia mammillata, we first built a high-resolution atlas of embryonic cell lineages, cell shapes, and cell interactions. We imaged 10 live embryos every 2 min up to the end of the neurula stages using multiview light-sheet microscopy. To systematically measure the developmental variability of a range of temporal and spatial cellular features, we developed a robust and scalable adaptive segmentation and tracking of embryonic cells procedure (ASTEC) compatible with high-throughput multiview light-sheet imaging datasets. We related these features to cell fate specification, which in ascidians is mainly controlled by differential sister cell inductions. Inspired by previous work indicating that the area of contact to signaling cells controls ascidian neural induction, we integrated our geometric description with a signaling gene expression atlas. This integration allowed us to test, through computational and experimental approaches, the hypothesis that contact area–dependent cell communication imposes constraints on embryonic geometries.

RESULTS

We found that, up to the neurula stages, Phallusia embryos develop without cell growth, programmed cell death, or cell neighbor exchanges. Beyond cell position, cell cycle duration, and cell lineages, we observed a high reproducibility of cell arrangements: 75% of cells shared at least 80% of their neighbors in all 10 embryos studied. Furthermore, the areas of contact between homologous cells varied by less than 20% across embryos. Mechanistically, we uncovered a tight link between the control of cell arrangements and asymmetric cell divisions, which give rise to sister cells of distinct fates. We then combined computational and experimental approaches to reveal that areas of cell contact between signaling and responding cells have sufficient encoding potential to explain all known early embryonic inductions, without the need to invoke gradients of ligand concentration. Finally, using geometrical perturbations of embryonic development we demonstrated that precise areas of cell-cell contact were important for mesendodermal and neural fate specification.

CONCLUSION

Our work establishes the highly reproducible ascidian embryo as a framework to bridge cell behaviors, morphogenesis, and the underlying regulatory program. The ASTEC pipeline allows systematic automated whole-cell segmentation and tracking across whole embryos in high-throughput light-sheet datasets. Second, we establish the geometric control of embryonic inductions as an alternative to classical morphogen gradients and suggest that the range of cell signaling events sets the scale at which embryonic reproducibility is observed. Finally, our study suggests that the high level of reproducibility of ascidian embryonic geometries may paradoxically lift constraints on the evolution of ascidian genomes, thereby contributing to rapid molecular evolution.

Reconstruction and modeling of Phallusia embryogenesis.

(Left) Quantitative analysis of Phallusia embryogenesis. We combined live light-sheet imaging of cell membranes (left images) with automated cell segmentation and tracking with color-coded cell fates (center images) to extract quantitative cell morphological properties (right images, color-coded by cell compactness). From top to bottom: embryo at the 64-cell, mid-gastrula, and late gastrula stages. (Right) Cell signaling model. We first made simplifying assumptions concerning the distribution and diffusion of signaling pathway components (top) and then integrated cell contact geometry with gene expression profiles to predict pathway activation levels in single cells (center) and binarized induction status (bottom).

Abstract

Marine invertebrate ascidians display embryonic reproducibility: Their early embryonic cell lineages are considered invariant and are conserved between distantly related species, despite rapid genomic divergence. Here, we address the drivers of this reproducibility. We used light-sheet imaging and automated cell segmentation and tracking procedures to systematically quantify the behavior of individual cells every 2 minutes during Phallusia mammillata embryogenesis. Interindividual reproducibility was observed down to the area of individual cell contacts. We found tight links between the reproducibility of embryonic geometries and asymmetric cell divisions, controlled by differential sister cell inductions. We combined modeling and experimental manipulations to show that the area of contact between signaling and responding cells is a key determinant of cell communication. Our work establishes the geometric control of embryonic inductions as an alternative to classical morphogen gradients and suggests that the range of cell signaling sets the scale at which embryonic reproducibility is observed.

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