ABSTRACT: Collective behaviours are key in development. In a cell population, cells must coordinate to take on the right identity at the right place and time. In an embryo, cell populations must coordinate to undergo robust patterning and morphogenesis. The mechanisms underlying these coordinated behaviours remain poorly understood, in part because answering these questions requires the extensive bridging across biological scales—from molecules to whole embryos. In this thesis, I address these questions across two projects both using the chick embryo as a model system. Chapters 3-5 (“collective cell invasion project”; PhD years 1-3) investigates the first question, “How do cells coordinate to take on the right identity at the right place and time?” through examining how morphogen signalling, cell fate specification, and epithelial-to-mesenchymal transition (EMT) are coordinated during two contexts of collective cell invasion. First, the ingression and subsequent migration of nascent mesoderm through the morphogenetic structure called the primitive streak during gastrulation. Second, the delamination and subsequent migration of neural crest cells from the dorsal neural tube during neurulation. By comparing across developmental contexts and utilising a mixture of classical and modern techniques (experimental embryology, gain-of-function and loss-of-function assays, transcriptomics), I show that EMT and bone morphogenetic protein (BMP) signalling are reciprocally coupled to drive collective cell invasion. This coupling provides a molecular mechanism for synchronising cell fate specification with invasion progression, repositioning EMT as not just a process that guides changes in cell behaviour but as an active regulatory process that modulates morphogen signalling activity to coordinate tissue-level behaviours during collective cell invasion. Chapters 6-7 (“developmental checkpoint project”; PhD year 4 onwards) investigates the second question, “How do embryonic cell populations coordinate to undergo robust patterning and morphogenesis?” through the identification of a novel developmental bottleneck that gates the transition between primary gastrulation and axis elongation. Convergent evidence from several orthogonal approaches (tracking of normal developmental progression, resilience to temperature-induced diapause, regulative capacity following node ablation, and sensitivity to toxins) all point to the existence of a global embryonic bottleneck at Hamburger and Hamilton (HH) stage 4+. Further analysis reveals that this HH4 to HH5 transition seems to be dependent on the level of translation, providing a mechanistic basis for understanding coordinated developmental progression. Bulk RNA sequencing experiments further shed light on potential mechanisms that may be regulating the HH4 to HH5 transition, including surveilling translation activity. Future work will test whether the mechanisms underlying this novel bottleneck satisfies the four criteria of a checkpoint: dependency, active enforcement, separability, and dispensability. Together, these two projects demonstrate that collective behaviours in the embryo are achieved in part by distinct mechanisms—molecular coupling between morphogen signalling and EMT at the level of invading cell populations and translational regulation gating a global transition at the level of the whole embryo—thus advancing our understanding of how development is coordinated across different biological scales.
