Primordial germ cells (PGCs) are the precursors of gametes, and a presumptive PGC-like cell (PGCLC) population was recently identified in mouse gastruloids. A new paper in Development investigates this population in detail and offers new insight into the signals that contribute to PGC specification and maturation. To hear more about the story, we caught up with first author Christopher Cooke and corresponding author Naomi Moris, Group Leader at the Francis Crick Institute, UK.

Chris (right) celebrating in the pub on the day of his PhD viva with his supervisor, Naomi (left), and external examiner, Ramiro Alberio (centre).

Naomi, can you give us your scientific biography and the questions your lab is trying to answer?

NM: I did a PhD on the epigenetic control of cell fate decisions with Cristina Pina and Alfonso Martinez Arias at the University of Cambridge, UK, then stayed on with Alfonso to look at how these decisions are coordinated using 3D gastruloids, which were first developed in his lab, and eventually got a Junior Research Fellowship at Newnham College to translate the mouse gastruloid protocol to humans. In 2021, I started my own group at the Francis Crick Institute, where we're bringing all of that together to try to use a range of embryo-like models to answer fundamental questions in developmental biology. I would say that we have two main themes in the lab: one using human gastruloids and extended models to develop new tools and, most importantly, to begin to answer questions around human development, while the other theme revolves around an interest in germ cell biology, for which we mostly use mouse gastruloids.

Christopher, how did you come to work with Naomi and what drives your research today?

CC: I first met Naomi along with Alfonso when they came to discuss gastruloids at Abcam. As it happens, my team manager and I were looking for a suitable PhD opportunity for me and luckily both Alfonso and Naomi were willing to take me on as a PhD student. I was very fortunate to get to work with them both in the dynamic, collaborative environment they had cultivated and in such an exciting field of research. During my PhD, Naomi encouraged me to investigate potential PGCLCs in the gastruloids and we continued to work on this project throughout the pandemic, even after Naomi had started her own lab at the Crick. When Alfonso moved his lab to Barcelona towards the end of my PhD, Naomi kindly invited me to work with her and her lab in the Crick, which was too great an opportunity to turn down! I'm now back at Abcam so my work is more industry focused, which offers somewhat different challenges and rewards compared with academic research.

Before your work, what was known about PGC development, and what sort of in vitro models were available?

CC: A fair amount was known from the mouse embryo, including where these cells form, the environment required for their formation and how they progress over time. However, several questions still remained; for instance, what controls the exact number of PGCs that form? Studying PGCs in the mouse embryo is not an easy task as they first appear post-implantation and start as such a small population. So, in vitro models offer ways to circumvent some of these issues.

PGCLCs can be generated from several in vitro models, usually using cells exposed to a cocktail of signalling factors to generate high numbers of PGCLCs. This method allows scientists to gain a much greater understanding of the genetic/epigenetic mechanisms at play in PGCLCs, but it doesn't allow them to be studied alongside other cell types that they would normally interact with within the embryo.

Can you give us the key results of the paper in a paragraph?

CC: We demonstrate that a population of PGCLCs consistently form within mouse gastruloids and display many of the characteristic features of their in vivo counterparts. This includes interactions with endodermal cells and migratory behaviour. Interestingly, we show that these gastruloid-derived PGCLCs can reach a developmental stage much later than do PGCLCs derived from embryoid bodies, without the need for addition of gonadal cells. Finally, by studying various signalling pathways associated with PGCs, we found that BMP signalling had little effect on the formation of gastruloid-derived PGCLCs, but FGF inhibition led to a significant increase, in a concentration-dependent manner. These findings show that gastruloids offer a suitable system in which to study the development and maturation of PGCLCs.

Were you surprised that BMP signalling appeared to have little effect on PGCLC specification in the gastruloids?

CC: This was definitely quite a surprising finding as it was widely considered a ‘given’ that BMP4 signalling is required for PGC specification, and the current in vitro method of deriving PGCLCs relies on BMP treatment. But, some of the literature does suggest the possibility that BMP may not be directly responsible for PGC specification and some recent findings by other groups had also hinted at a less vital role for BMP. Then, shortly after we had seen this in the gastruloids, Sophie Morgani and Kat Hadjantonakis reported similar findings in the micropattern system (Morgani and Hadjantonakis, 2021), which helped us feel more assured in our interpretation of our results.

Gastruloid-derived PGC-like cells within the endodermal tract of a mouse gastruloid. Grey, Hoechst; cyan, Blimp1:GFP; yellow, AP2g; magenta, FOXA2. Image credit: Christopher Cooke.

Gastruloid-derived PGC-like cells within the endodermal tract of a mouse gastruloid. Grey, Hoechst; cyan, Blimp1:GFP; yellow, AP2g; magenta, FOXA2. Image credit: Christopher Cooke.

What are the implications of this work for future in vitro studies of lineage specification?

NM: I think beyond the fact that gastruloids can produce PGCLCs, it's most remarkable that they reach a relatively mature stage of development just with spontaneous ‘co-development’ between the tissues and cell types present in gastruloids. That has implications even outside the PGC field, because rather than searching for protocols that give us the purest populations (as is often done in settings for which particular outcomes are required, such as for applied research), it teaches us that by tapping into the innate potential of pluripotent cells, and exposing them to conditions similar to that of the early embryo, we can potentially generate cell types of interest that might otherwise be hard to derive in the lab. That diversity of cell types, working together in an organised way, is actually incredibly powerful.

I think beyond the fact that gastruloids can produce PGCLCs, it's most remarkable that they reach a relatively mature stage of development just with spontaneous ‘co-development’ between the tissues and cell types present in gastruloids

When doing the research, did you have any particular result or eureka moment that has stuck with you?

CC: I think I was quite lucky with this research to have a few results and findings which have stayed with me, making all the late nights in the labs and working at weekends worthwhile. Seeing the effect that FGF inhibition had on the PGCLC population, finding clear signs of PGCLC maturation within the gastruloids, plus a few other things come to mind. I found the malleability of the gastruloid system was definitely a high point, as I enjoyed the ability to modulate signalling pathways and study how that impacts various cell populations within the system in a predictable manner.

NM: For me, it was when Chris came to show me the first two-photon, live-imaging shots of the PGCLCs migrating. He'd been battling to try and see them with confocal microscopy and several other methods, but they are so internalised in the gastruloid and the cells are so sensitive to light that it was very difficult to get clear live movies. So, when we saw the first tantalising evidence that they might be moving, and possibly also directionally, it was wonderful and I was so proud of him for persevering to get that piece of the puzzle!

And what about the flipside: any moments of frustration or despair?

CC: Surprisingly, I think overall there were no real low points, at least that I can think of now. There was the inevitable self-doubt, and it was initially challenging to prove to myself that the cells were definitely PGCLCs, because PGCs and pluripotent cells express a lot of the same proteins. Also, undertaking a PhD through the pandemic came with plenty of challenges, but also gave me the opportunity to take a step back, assess data and re-focus efforts. Overall, I feel very lucky to have been able to study PGCLCs in the gastruloid system.

Christopher, what is next for you after this paper?

CC: As part of the funding agreement with Abcam I have returned to work at the company post-PhD. I'm currently working in the Cell Engineering team where I'm looking after custom gene-editing projects in Cambridge. It's all fairly new to me, but I'm always happy to learn new skills and techniques, especially when they have so many useful applications. Ideally, I hope to combine my PhD experience in stem cell and developmental biology with my newly acquired gene-editing skills in the not-too-distant future.

Naomi, where will this story take your lab next?

NM: We're continuing this line of research, figuring out exactly how mouse gastruloids are able to specify PGCLCs and promote their maturation, and whether, not only the transcriptional and protein-level characterisation of these cells matches that of the embryo, but indeed whether their epigenetic characteristics might also show interesting maturation, because that's crucial for them to be functionally capable of giving rise to gametes.

Finally, let's move outside the lab – what do you like to do in your spare time?

CC: I try to take any opportunity to get out in nature, whether hiking in the UK, abroad on solo trips around Europe or much closer to home walking around Cambridge. I also enjoy playing games with friends and family, whether card, board or computer based. During my PhD, I was responsible for starting a Pub Assessment society within the lab, so I like to continue that vital work whenever I can.

NM: Since recently stepping back from tissue culture to focus on running the lab, I've felt a need to fill that void and now enjoy growing all sorts of weird and wonderful plants and flowers from seed – popping each one into individual wells of a seed tray and hoping for germination reminds me of a 96-well plate and waiting for gastruloid elongation, although I still have my fair share of failures so I guess I have a lot to learn!

C.C. & N.M.: Francis Crick Institute, London NW1 1AT, UK.C.C.: Department of Genetics, University of Cambridge, Cambridge, UK and Abcam, Cambridge Biomedical Campus, Cambridge, UK.

E-mail: [email protected]

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