Germ granules have been hypothesized to deliver mRNAs of germ cell fate determinants to primordial germ cells. Now, a new study in Development finds that many mRNAs enriched in germ granules are not involved in germline development in Caenorhabditis elegans. To find out more about the story behind the paper, we caught up with first author Alyshia Scholl, second author Yihong Liu and corresponding author Geraldine Seydoux, Professor at Johns Hopkins University School of Medicine.
Alyshia Scholl (left), Geraldine Seydoux (centre) and Yihong Liu (right)
Geraldine, can you give us your scientific biography and the questions your lab is trying to answer?
GS: I was ‘born and raised’ a C. elegans developmental geneticist having done both my PhD and postdoc in the field. I was drawn to germ granules early on when Susan Strome described their segregation to the embryonic germline in C. elegans. In 1999, my first postdoc K. Subramaniam showed that the Nanos homolog nos-2 localizes to germ granules and is required for primordial germ cells (PGCs) to generate the germline. Since then, I always had the hope of one day describing the full germ granule transcriptome. It was not until postdoc Sean Lee showed that the germ granule protein MEG-3 binds 500+ mRNAs that we could finally begin to characterize systematically transcripts in germ granules.
Alyshia and Yihong, how did you come to work in the lab?
AS: Before graduate school, I was interested in understanding how different cell types are specified during development. When I joined Johns Hopkins as a graduate student, I naturally gravitated towards Geraldine's lab in understanding how C. elegans specify their PGCs.
YL: I was looking to get into research as an undergraduate student and came across Geraldine's lab. After meeting with Geraldine and lab members, I felt that it would be a good fit for me and decided to join.
What is known about the mRNA transcripts enriched in germ granules before this work?
GS: Previously, it was known that germ granules contain polyadenylated mRNAs. A survey in Drosophila identified 58 transcripts, including Nanos, that appeared to localize to germ granules (Rangan et al., 2008). However, no one had systematically conducted a screen for germ granule RNAs. When the lab identified RNAs bound to MEG-3 (Lee et al., 2020), we used that list as the basis for a single molecule in situ fluorescent hybridization (smFISH) screen to identify germ granule transcripts.
I always had the hope of one day describing the full germ granule transcriptome
Can you give us the key results of the paper in a paragraph?
GS: Germ granules had been hypothesized to localize mRNAs required for germ cell fate specification. We were surprised to find that most germ granule transcripts in C. elegans encode general housekeeping functions and are degraded before the formation of the PGCs. Only a minority are maintained to the PGC stage and, among those, only a handful are required for germ cell fate.
Were you surprised to find that the majority of mRNA enriched in germ granules do not persist into PGCs and are not associated with germ cell fate specification?
GS: Yes! We were hoping to identify a long list of transcripts required for germ cell fate. Instead, we found only nine, all of which were already known to play a role in germ cell development. We did also identify 13 germ granule transcripts conserved between C. elegans and Drosophila, but again these were a small minority.
Airlocalize-processed image of the germ granule RNAs grif-1 (green) and nos-2 (magenta) in a four-cell embryo (maximum projection).
If most of the germ granule mRNAs are not essential for germ cell fate determination, what do you think is the role of these mRNA transcripts?
GS: From previous work, we knew that RNA enhances the ability of MEG-3 to form condensates that stabilize the protein-rich phase of P granules. Therefore, we hypothesize that most transcripts in P granules function as structural scaffolds to support MEG-3 condensation. Only a minority of transcripts (including nos-2) use the granules to enrich in the PGCs.
Alyshia and Yihong, when doing the research, did you have any particular frustrations or eureka moments that have stuck with you?
AS: The results of the in situ and RNAi screens were initially very disappointing as I was hoping to identify new genes required for germ cell fate. Another surprise was that the fate of germ granule transcripts does not seem to require the mRNAs to be inside of the granules: RNA degradation and translation proceed normally in meg-3 meg-4 mutants that fail to recruit mRNAs to granules. These results were not what I initially expected but showed me how data can take you in unexpected directions.
These results were not what I initially expected but showed me how data can take you in unexpected directions
YL: I developed a systematic pipeline to quantify the number of RNA molecules in granules. I was surprised to see how much quantifiable information can be extracted from in situ images! We found that RNAs in granules concentrate in clusters containing several copies of the same transcript, as first shown in Drosophila. The significance of these ‘homotypic clusters’ is still unclear and suggests that we still have much to learn about how mRNAs are stored in the cytoplasm.
Why did you choose to submit this paper to Development?
GS: Our work addresses an outstanding question in the field of developmental biology. Development is a journal run by developmental biologists for developmental biologists! We felt our work would reach the right audience. The review process at Development is also great and very much helped us improve the initial manuscript.
Alyshia and Yihong, what is next for you after this paper?
AS: I'm currently looking for postdoctoral fellowship positions where I can expand this work into the formation and maintenance of the germline stem cell niche.
YL: I will be starting graduate school in the fall to continue doing biological research.
Geraldine, where will this story take your lab next?
GS: We are still interested in figuring out the mechanisms that specify germ cell fate in embryos. The few germ granule mRNAs that are involved in this process give us a good starting point. Some of these transcripts enrich in the granules with high efficiency and we want to know how this specificity is achieved. We are also developing methods to sequence the first zygotic transcripts expressed by PGCs. We hope that this will help us understand better what shapes the maternal-to-zygotic transition in the nascent germline.
Finally, let's move outside the lab – what do you like to do in your spare time?
GS: Travel!! In fact, Yihong is traveling in the UK and China and I am on a hike in Cinque Terre, Italy. Alyshia will soon travel to Florida where she will present this work at the FASEB RNA Localization and Local Translation Conference. Travel is a source of rejuvenation for all of us!
HHMI and Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
E-mail: [email protected]
