Periodic Reporting for period 1 - JOLI (Using the jellyfish Clytia hemisphaerica to explore the first steps of meiosis by live-imaging.)
Reporting period: 2019-05-01 to 2021-04-30
The overall objective of this project was to provide a framework for investigating meiosis during oogenesis in Clytia by development of specific molecular tools and approaches to establish a spatiotemporal cartography of early meiotic events within the developing gonad. More specifically the aims were 1) To develop and deploy a variety of tools and methods to characterize early meiosis (in-situ hybridization for gene expression analysis and telomere detection, Clytia specific antibodies for key molecular actors of synapsis to allow their tracking by immunohistochemistry, vital dyes for live imaging), 2) To conduct pilot studies to explore possibilities live imaging of the Clytia gonad, and 3) to study of the function of spo11 in Clytia meiosis via the generation of CRISPR mutants.
In pioneering functional studies in this system, we used CRISPR-Cas9 knockout methods to generate several male and female jellyfish lines carrying mutations for Spo11, a key actor in the initiation of meiotic recombination whose precise role during cross species remains to be understood. The mutants showed several clear phenotypes: all spo11 mutants show a clear loss of recombination and pairing in fully grown prophase arrested oocytes, while the formation of the synaptonemal complex was fully or partially abolished. We developed antibodies against the rad-51 protein (optimization ongoing) to assess double stranded breaks. Further analysis showed that after fully grown mutant oocytes are triggered to undergo maturation, meiotic divisions are disrupted such that the resulting female gamete is diploid or tetraploid. Unexpectedly, these eggs can be successfully fertilised , and are capable of producing viable offspring.
These results have been disseminated via three scientific conferences either online or in person. Additionally, the project has been communicated to the broader public via science days and lectures. Finally, the results of this project will be written up in two scientific papers to be submitted in the next year, as well as two review papers (one published, another submitted). The data and mutant lines generated in this work will be of use for several future projects, now that Clytia is firmly established as an excellent model to investigate early as well as late events of meiosis.