A gastruloid-based model system to study human and non-human primate primordial germ cell specification and reprogramming

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Primordial germ cells (PGCs) are embryonic precursor cells, which give rise to sperm or eggs. PGCs specification starts during early embryonic development, concurrent with the beginning of gastrulation, at around week two in the human embryo post-implantation. Understanding how PGCs develop will provide exciting prospects for future medical applications and address key developmental questions. However, it is not possible to study PGCs using human embryos due to ethical concerns. Thus, I aim to make use of embryo-like in vitro models. These so-called gastruloids are generated by self-organization of pluripotent stem cells in three-dimensional aggregates, which thereby undergo early key developmental events. Gastruloids offer an unprecedented potential to study primate early embryogenesis due to their realistic cytoarchitecture, organization, and temporally controlled cell fate transitions. In addition, mouse gastruloids can give rise to PGC-like cells in a spatiotemporally organized location relative to somatic tissue-like structures. I aim to examine PGC specification and reprogramming in human and non-human primate (NHP) gastruloids. Using both human and macaque cells in parallel will open up the possibility to validate data obtained in vitro with available NHP in vivo data. Furthermore, comparative analysis will allow me to dissect potential similarities and differences between NHP and human PGC specification and early differentiation. After establishing and characterizing the primate-specific gastruloid system, I will perform conditional loss-of-function experiments to address the question: What is the role of pluripotency factors during PGC fate determination and reprogramming? For this, I will use an inducible degron system for rapid and comprehensive depletion of candidate transcription factors at different time points during gastruloid formation and evaluate the impact on the PGC population. Altogether, this project will constitute a defined in vitro system to study PGC specification and reprogramming in primates. The combined set of tools proposed in this application can be applied for drug toxicity testing, congenital disease modeling and contribute to the 3Rs (reduce, refine, and replace) in animal experimentation.