About the Project
Our research is focused on understanding the mechanisms and regulation of chromosome segregation in mitosis. During the cell cycle, accurate chromosome segregation ensures that both daughter cells inherit the correct complement of chromosomes. Errors in this process cause aneuploidy leading to cancer and developmental defects. Duplicated sister chromatids are segregated in mitosis by a large cellular apparatus, the bi-polar mitotic spindle. The spindle is organized from centrosomes, located at opposite poles of the cell. At metaphase, condensed sister chromatid pairs are aligned on the metaphase plate at the centre of the mitotic spindle. Each chromatid is attached to microtubules by kinetochores, large protein complexes that specifically assemble onto centromeric chromatin. Once all chromosomes achieve bi-polar orientation on the mitotic spindle, and tension is exerted at the kinetochore-microtubule attachment site, anaphase is triggered. This results in the loss of sister chromatid cohesion and the segregation of each sister chromatid to opposite poles of the cell, a process powered by microtubule depolymerization.
Our current research is directed to reconstituting mitosis in vitro to understand the structure and mechanism of the kinetochore and spindle pole body in budding yeast, how the SAC is activated, and how microtubules mediate chromosome movement in anaphase.
The PhD project will involve joining a team that is reconstituting mitosis in vitro in order to understand the structures of the kinetochore, how it attaches chromosomes to the mitotic spindle and how it activates the spindle assembly checkpoint in response to loss of attachment to microtubules and lack of tension (indicating loss of bipolar attachment). The project will include a variety of techniques including single particle cryo-electron microscopy, cryo-electron tomography, super-resolution fluorescence microscopy and in vitro reconstitution approaches.
References
Reviews
Alfieri, C., Zhang, S. and Barford, D. Visualizing the complex functions and mechanisms of the anaphase-promoting complex/cyclosome (APC/C). Open Biol. (review). pii: 170204. doi: 10.1098/rsob.170204 (2017).
Biggins, S. The composition, functions, and regulation of the budding yeast kinetochore. Genetics 194, 817-846, doi:10.1534/genetics.112.145276 (2013).
Westermann, S. & Schleiffer, A. Family matters: structural and functional conservation of centromere-associated proteins from yeast to humans. Trends in cell biology 23, 260-269, doi:10.1016/j.tcb.2013.01.010 (2013).
Cheeseman, I. M. The kinetochore. Cold Spring Harbor perspectives in biology 6, a015826, doi:10.1101/cshperspect.a015826 (2014).
Musacchio, A. & Desai, A. A Molecular View of Kinetochore Assembly and Function. Biology 6, doi:10.3390/biology6010005 (2017).
Hinshaw, S. M. & Harrison, S. C. Kinetochore Function from the Bottom Up. Trends in cell biology 28, 22-33, doi:10.1016/j.tcb.2017.09.002 (2018).
Primary Literature
Yan, K., Yang, J., Zhang, Z., McLaughlin, S.H. and Barford, D. Architecture of the kinetochore CBF3-CEN3 DNA complex of budding yeast. Nature Structural Molecular Biology, 25,1103-1110 (2018).
Alfieri, C., Chang, L., Zhang, Z., Yang, Y., Maslen, S., Skehel, M. and Barford, D. Molecular basis of APC/C regulation by the spindle assembly checkpoint. Nature, 536, 431-436 (2016).
Zhang, S., Chang, L., Alfieri, C., Yang, J., Zhang, Z., Maslen, S., Skehel, M. and Barford, D. Molecular mechanism of APC/C activation by mitotic phosphorylation. Nature, 533, 260-264 (2016).
Chang, L., Zhang, Z., Yang, J., McLaughlin, S.H. and Barford, D. Atomic structure of the APC/C and its mechanism of protein ubiquitination
Nature, 522, 450-454 (2015).
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