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Single-cell genomics in rabbit and mouse elucidate eutherian embryonic development



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Ton, Mai-Linh 


Biomedical research relies heavily on the use of model organisms to gain insight into human health and development. Traditionally, the mouse has been the favoured vertebrate model, due to its experimental and genetic tractability. Non-rodent embryological studies however highlight that many aspects of early mouse development, including the egg-cylinder topology of the embryo and its method of implantation, diverge from other mammals, thus complicating inferences about human development.

To get a better understanding of rabbit development, we constructed a morphological and molecular atlas of rabbit development, which like the human embryo, develops as a flat-bilaminar disc. We report transcriptional and chromatin accessibility profiles of almost 180,000 single cells and high-resolution histology sections from embryos spanning gastrulation, implantation, amniogenesis, and early organogenesis. Using a novel computational pipeline, we compare the transcriptional landscape of rabbit and mouse at the scale of the entire organism, revealing that extra-embryonic tissues, as well as gut and primordial germ cell (PGC) cell types, are highly divergent between species.

Focusing on these extra-embryonic tissues, which are highly accessible in the rabbit, we characterize the gene regulatory programs underlying trophoblast differentiation and identify novel signalling interactions involving the yolk sac mesothelium during haematopoiesis. Finally, we demonstrate how the combination of both rabbit and mouse atlases can be leveraged to extract new biological insights from sparse macaque and human data. The datasets and analysis pipelines reported here set a framework for a broader cross-species approach to decipher early mammalian development, and are readily adaptable to deploy single-cell comparative genomics more broadly across biomedical research.

Due to the genetic tractability of mouse, we aimed to interrogate the role key transcription factors (TFs) such as Zic2/3, Mix-like 1(Mixl1), Eomesodermin (Eomes), Stat3, and Runx1 in early development. These key TFs are involved in a variety of roles, such as metabolism, as well as the fate decision of mesoderm, and blood development. Constructing an understanding of the stepwise role that these TFs play in sequence for developing blood and mesoderm allows for a better understanding of the consequences of genetic mutations. To perform this analysis, we generate a series of clustered regularly interspaced short palindromic repeats (CRISPR)-mediated knock out cell lines. Using a chimaera model system where knock-out (KO) cells are injected into wild-type (WT) host blastocysts, we can understand the cell-autonomous role that each TF plays.

In conclusion, studying single-cell transcriptomics unveils the molecular profiles behind each cell type; however combining with spatial information, chromatin accessibility, and gene perturbations allows for further understanding of the signalling niche and regulatory elements behind cell type diversification. Augmenting this analysis by a variety of model organisms allowed for a nuanced understanding of eutherian development, such as macaque and human development by understanding the divergent and convergent features of embryonic development. This also allows for the optimisation of in vitro differentiation protocols in the future. Additionally, this has implications for biomedical research due to the species- and cell-type-specific responses to drug screens. A comprehensive understanding of each target cell type of interest allows for researchers to better adapt model systems to their intended target.





Gottgens, Berthold


10x genomics, comparative embryology, crispr, embryology, Eomes, fate choice, gastrulation, haematology, hematology, macaque, Mixl1, mouse embryo, rabbit, Runx1, scATAC-seq, scRNA-seq, single cell, single cell genomics, single cell multiome, Stat3, stem cell biology, Zic2, Zic3


Doctor of Philosophy (PhD)

Awarding Institution

University of Cambridge
Harvard Herchel Smith PhD Fund.