Before fertilisation, oocytes arrest in metaphase in humans, as well as many other animal species including mouse and Drosophila. This arrest lasts for a long time, 12-24 hours in humans. During this arrest, oocytes need to maintain the bipolar spindle for accurate chromosome segregation. A failure to maintain the bipolar spindle in oocytes could result in aneuploidy in eggs, which leads to miscarriages or congenital condition such as Down syndrome in human. Despite the potential medical implication, how oocytes maintain the bipolar spindle during this arrest is not clear. Recently the lead supervisor’s lab found that multiple spindle proteins change their localisation during this arrest in Drosophila oocytes, and that these changes are important for stably maintaining spindle bipolarity during the arrest (Costa & Ohkura, 2019). The molecular basis of these changes is unknown, and limited availability of materials is a challenge to research into oocytes.
(1) to establish single-cell proteomics in oocytes to identify molecular changes during metaphase arrest.
(2) to test whether these molecular changes are conserved.
(3) to define the roles of these changes in accurate chromosome segregation and fertility in oocytes.
Methodology: A method for single-cell quantitative proteomics will be developed for Drosophila and mouse oocytes to identify the changes of protein levels during metaphase arrest. In addition, manual staging and pooling of a small number of oocytes will allow mass-spectrometry to identify changes in post-translational modifications in oocytes. The data will be analysed and compared between the species to select proteins for further investigation. To test the biological significance of these changes, fertility, accuracy of chromosome segregation, protein localisation, spindle bipolarity and chromosome-spindle interaction will be assessed after interfering with these changes in oocytes.
Through PhD study, the student will develop into an independent researcher who can tackle a biological question using multiple approaches, including microscopy, image analysis, genetics, biochemistry, proteomics and bioinformatics. The student will gain project management skills by directing the project through interactions with researchers in multiple labs and institutions. Training for microscopy, genetics and molecular biology will be provided mainly by the lead supervisor in Edinburgh. Training of quantitative mass-spectrometry and computational 4D imaging analysis will be provided by the second and external supervisors in the Wellcome Centre for Cell Biology in Edinburgh and Karolinska Institute in Stockholm (Ly et al., 2017; Kouznetsova et al, 2019). Single-cell quantitative proteomics using oocytes will give the student an opportunity of technology development by combining optimisations in experimental steps on the bench and data acquisition/analysis in silico. Training for statistics and programming will be provided through workshops and on-job training. Regular one-to-one training for various writing and oral/poster presentations will be provided by the lead supervisor throughout the PhD study.
This MRC programme is joint between the Universities of Edinburgh and Glasgow. You will be registered at the host institution of the primary supervisor detailed in your project selection.
All applications should be made via the University of Edinburgh, irrespective of project location. For those applying to a University of Glasgow project, your application along with any supporting documents will be shared with University of Glasgow. http://www.ed.ac.uk/studying/postgraduate/degrees/index.php?r=site/view&id=919
Please note, you must apply to one of the projects and you must contact the primary supervisor prior to making your application. Additional information on the application process is available from the link above.
For more information about Precision Medicine visit: http://www.ed.ac.uk/usher/precision-medicine
1. Costa MFA. and Ohkura H (2019) The molecular architecture of the meiotic spindle is remodeled during metaphase arrest in oocytes. J. Cell Biol. 218: 2854-2864.
2. Ly T, Whigham A, Clarke R, Brenes-Murillo AJ, Estes B, Madhessian D, Lundberg E, Wadsworth P, Lamond AI (2017) Proteomic analysis of cell cycle progression in asynchronous cultures, including mitotic subphases, using PRIMMUS. Elife. 6: e27574.
3. Kouznetsova A, Kitajima TS, Brismar H, Höög C (2019) Post-metaphase correction of aberrant kinetochore-microtubule attachments in mammalian eggs. EMBO Rep. 20: e47905.