Analysing cardiac specification and morphogenesis using mouse embryoids

Background:

This project will address how embryonic cells develop into distinct cardiac lineages and form a heart during morphogenesis. Addressing this question has important implications for our understanding of congenital heart diseases -affecting ~1% of newborns- and the design of regenerative methods targeting specific populations of heart cells.

Our heart comprises four chambers, two ventricles and two atria, driving blood from the body to the lungs and through the rest of the body. We recently established a fate map demonstrating ventricular and atrial cardiac progenitors have distinct spatial and temporal origins in the primitive streak (the early embryonic structure giving rise to mesodermal and endodermal tissues) (Ivanovitch et al., 2021). We further demonstrated that the primitive streak cells contributing to the ventricles have a distinct molecular signature from those forming the atria. These observations suggest cardiac progenitors are already prespecified in the streak; and this may prefigure their allocation to distinct anatomical structures of the heart.

The student will exploit a 3D mouse stem cell-based model resembling natural embryos (embryoids) developed by Prof. Zernicka-Goetz's team and Colleagues to address this question (Amadei et al.,2022, Lau et al., 2022, Tarazi et al. 2022). The embryoids are remarkably like natural embryos. They develop a similar morphology, with a beating heart tube, blood circulation, a brain and spinal cord and the beginnings of a gut tube. They may present a unique opportunity to study the molecular mechanisms regulating cell fate decisions and morphogenesis in vitro as an alternative/complement to in vivo experiments.

Objectives and methods
 

The project aim is to uncover the mechanisms underlying the specification and morphogenesis of chamber-specific cardiomyocytes. We recently found that cardiac progenitors are already assigned to specific heart compartments in the primitive streak. Anterior regions of the primitive streak -expressing low levels of the T-box transcription factor T- contribute to the ventricles. In contrast, the posterior primitive streak -expressing high levels of T- contributes to the atria.

1) The student will test the hypothesis that ventricular or atrial cardiomyocytes (transcriptionally and functionally distinct) specification depends on initial molecular conditions set in the primitive streak.

They will first generate distinct populations of primitive streak-like cells expressing low or high levels of T using mES cells in 2D culture. They will test their ventricular and atrial differentiation potential. In the low T condition, they will then overexpress T using an inducible overexpressing cell line (TTET-ON line) and analyse the putative changes in differentiation potential induced by T overexpression. They may extend the analysis to other genes differentially expressed in the Tlow versus Thigh primitive streak.

2) They will analysed the cardiac lineages’ emergence from the primitive streak in the embryoids in collaboration with Prof. Zernicka-Goetz's team. The embryoids undergo morphogenesis forming a beating looped heart as it occurs in natural embryos. They will assess reproducibility and similarities with natural embryos in detail.

3) Using the synthetic embryos, they will test if overexpression of transcription factors in the primitive streak (e.g. T or others) can modulate the proportions of ventricular vs atrial cardiomyocytes and consequently disrupt heart morphogenesis. Therefore, the student will implement embryoid as a novel tool for studying the molecular and genetic basis of cardiac lineage specification, morphogenesis and congenital heart diseases.

For this project, the student will work under the supervision of Dr Ivanovitch (project lead; ivanovitch-lab.com) and collaborate closely with Prof Magdalena Zernicka-Goetz's team at the University of Cambridge (zernickagoetzlab.com). You can read a press report on embryoids here: https://www.bbc.co.uk/news/health-62679322.

References:

1. Ivanovitch K et al. Ventricular, atrial and outflow tract heart progenitors arise from spatially and molecularly distinct regions of the primitive streak. PLOS Biology doi.org/10.1371/journal.pbio.3001200

2. Amadei G,. et al. Synthetic embryos complete gastrulation to neurulation and organogenesis. Nature. 2022 Aug 25. doi: 10.1038/s41586-022-05246-3.

3. Lau K.Y.C. et al. Mouse embryo model derived exclusively from embryonic stem cells undergoes neurulation and heart development, Cell Stem Cell, 2022, https://doi.org/10.1016/j.stem.2022.08.013.

4. Tarazi S. et al. Post-gastrulation synthetic embryos generated ex utero from mouse naive ESCs,

Cell, 2022, https://doi.org/10.1016/j.cell.2022.07.028.