University of Cambridge
Nodal organized cell dynamics for building tissue shapes in vertebrate gastrulation
Abstract
dc:description.abstractDuring development, complex tissue shapes form as a result of large-scale collective cell changes. How cell dynamics and mechanics link molecular scale genetic changes to tissue scale developmental outcomes is a major unresolved question in biology. Nodal is a key developmental regulatory gene from the TGF-β family that induces mesoderm and controls cell movement during gastrulation. How Nodal alters cell dynamics and mechanics is not well understood. To explore Nodal’s morphogenetic role in detail, we injected Nodal-related-2 (ndr2, cyclops) mRNA into one blastomere at the 128-cell stage animal pole of the zebrafish embryo, where Nodal is not normally expressed. The presence of Nodal+ cells led to a thickening of the animal pole blastoderm during the blastula stage. At the onset of gastrulation, the central region of the thickening internalizes, forming an ectopic blastopore with a circular, radially symmetrical blastopore lip, resembling a ‘volcano’ shape [1]. This tissue behaviour mimics the internalization process of the normal blastopore but takes place in the absence of other signalling pathways in this ectopic context, offering a simple model to study Nodal-organized cell dynamics. Next, I built an AI-based analysis pipeline for reconstructing 3D cells from confocal live imaging datasets to measure cell shapes and movements. Nodal injected cells are found to have larger cell volume and cell surface area, and to be more elongated in the z-axis (surface to deep from the animal pole view). They also move in a spatiotemporally patterned manner to form the volcano shape. To test the mechanical changes of the tissue as a result of these cellular changes, I measured cell membrane tension and tissue rigidity of the blastoderm thickening by Fluorescence Lifetime Imaging (FLIM) and Tissue Force Microscopy (TiFM). In conjunction with data from genetic perturbation experiments of p-stat3, Wnt/PCP, PI3K signalling performed by my collaborator, I constructed a full description of Nodal-mediated blastopore morphogenesis. First, the overlaying enveloping cell-layer of the Nodal expressing area lowers tension allowing the thickening to initiate, then cells undergo radial intercalation to move towards the animal pole while the spreading of these cells are blocked. The accumulation of cells at the animal pole, along with size and cohesion changes in these cells, cause cell jamming, leading to a higher pressure in this part of the blastoderm to continuously promote further thickening. Furthermore, cells with the highest level of Nodal perform internalization and pull surrounding cells to follow them, eventually creating the volcano/blastopore. Further, to explore functional similarities and differences of Nodal between zebrafish and chick during gastrulation, I implanted Nodal-coated Heparin beads at the area pellucida edge away from primitive streak in chick embryos during late HH2 or HH3 stages. I found that cells exposed to Nodal cluster along the dorsal-ventral axis and show ectopic Brachyury (a mesoderm marker) expression, and some treated embryos show a secondary body axis. These results suggest that Nodal may alter cell dynamics in a similar fashion to zebrafish as it drives gastrulation in chick. Overall, my project provides mechanistic insights on how Nodal signalling controls cell properties, collective migration, and tissue mechanics during gastrulation.
Degree
thesis:*- Name dc:type.qualificationname
- Doctor of Philosophy (PhD)
- Level dc:type.qualificationlevel
- Doctoral
- Grantor dc:publisher.institution
- University of Cambridge
- Year dc:date.issued
- 2024
Author and committee
dc:creator, dc:contributor.*- Author dc:creator
-
- Lan, Yisha
- Advisor dc:contributor.advisor
-
- Xiong, Fengzhu
Subjects
dc:subject × 5Rights
dc:rightsIdentifiers
dc:identifier.*- DOI dc:identifier.doi
- https://doi.org/10.17863/CAM.116748
- OAI identifier oai:identifier
- oai:www.repository.cam.ac.uk:1810/381628