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Report

Teaser, summary, work performed and final results

Periodic Reporting for period 1 - ACENTSPINTUB (Setting up the spindle in the mammalian egg for meiosis and embryo development)

Teaser

Summary

â€¢ Meiosis in mammalian oocytes is the process that generates haploid eggs able to be fertilized and support embryonic development. Oocyte meiosis is notoriously prone to mistakes leading to miscarriage, abortions or offspring with development disabilities. One of the most critical steps in meiosis is formation of a bipolar spindle that provides the machinery for accurate chromosome segregation. In mitotic cells, an organelle known as the centrosome plays a key role in organising microtubules to form the two spindle poles. By contrast, mammalian oocytes and early stage embryos lack classic centrosomes. Consequently, the spindle assembly depends on the self-organization of numerous microtubule organizing centres (MTOCs). Therefore, our main goals were 1) understanding how MTOCs are clustered to regulate bipolar spindle formation in acentriolar oocytes, and 2) understanding the functional properties of cytoplasmic MTOCs.
Conclusions:
Scientific achievements:
â€¢ We believe that we successfully achieved these two main goals. First goal: We have been the first to show that MTOC clustering in oocytes depends not only on microtubules, as previously thought, but also on F-actin. Moreover, we have found that F-actin recruitment to the spindle by Myosin X is required to carry out this function. these findings were presented last year (July 2018) at the annual meeting of the Society for the Study of Reproduction (New Orleans, USA). The EU was acknowledged in the abstract. It is of note, that this meeting is the worldâ€™s largest and most prestigious conference in the field Reproductive Biology. Moreover, this research was selected to receive the Lalor Foundation Merit award from the Society for the Study of Reproduction. This award recognizes the most meritorious and excellent research presented at the meeting. These results are currently being written up for submission to a top-tier journal as an open-access article and the EU will be acknowledged. Second goal: We have also been the first to unveil the function of cytoplasmic MTOCs in oocytes. We found that cytoplasmic MTOCs are necessary for anchoring polar MTOCs to the cortex. Moreover, such a connection of the spindle to the cortex is necessary to regulate spindle positioning. These results were accepted to be presented at the annual meeting of the Society for the Study of Reproduction (California, July 2019). The EU is acknowledged in the submitted abstract. Moreover, these results are currently being prepared for submission to a top-tier journal as an open-access article and the EU will be acknowledged.
Career achievements:
â€¢ In addition to achieving the research goals, Dr. Balboula received huge career training. The research conducted with this support increased Dr. Balboulaâ€™s scientific experience in molecular cell biology and gaining new experience in the fundamentals of cell division. For example, because Professor Gloverâ€™s lab is expert in imaging live cells, Dr. Balboula had the chance to successfully learn and use these methods to gain insight into how the acentriolar spindle is formed and how perturbations in signaling networks alter this process. This training was beneficial for Dr. Balboula, allowing him to establish an imaging system in his own laboratory and to continue studying oocyte and embryo biology while using new, emerging technologies. Moreover, Dr. Balboula participated in many career development programs such as Emerging Research Leaders\' Development Programme (ERLDP)â€.
Over all, the training which was provided to Dr. Balboula was critical in achieving his research goals and helped him to attain an independent faculty position at the University of Missouri, USA following completion of the Marie Curie Fellowship.

Work performed

We investigated the involvement of F-actin, a major cytoskeleton protein, in MTOC sorting and clustering in mouse oocytes using immunocytochemistry and time-lapse confocal microscopy. We found that MTOC clustering and sorting in oocytes depends not only on microtubules, as previously thought, but also on F-actin. Moreover, we found that F-actin recruitment to the spindle by Myosin X is required to carry out this function.
We also studied the biological significance of cytoplasmic MTOCs in mammalian oocytes. Our preliminary results revealed that cytoplasmic MTOCs are necessary for anchoring polar MTOCs to the cortex. Moreover, such a connection of the spindle to the cortex is necessary to regulate spindle positioning. These results were presented at international scientific conferences with a large number of audience and will be submitted for publication to appropriate journals.

Final results

The obtained results increased our understanding of two mysterious questions; 1) how MTOCs are clustered and sorted to form the spindle, and 2) what is the function of cytoplasmic MTOCs. Understanding the answer for these two questions is critical to understand why mammalian oocytes and embryos are error prone. Therefore, we believe that these results will be of interest not only to the basic research but also to infertility and IVF clinics.