The role of centrosomes in cell proliferation, division, and development
Final Report Abstract
Centrosomes are the major microtubule organizing centers in animal cells. The best understood function of centrosomes is to nucleate microtubules in order to catalyze the formation of a bipolar mitotic spindle, which is the molecular machine that ensures that both daughter cells inherit a proper genomic complement from the mother cell. This information is stored on chromosomes, which are segregated by the mitotic spindle to the two daughter cells during mitosis. Centrosomes were discovered over 130 years ago, however, our molecular understanding of centrosome function and dynamics are still limited. Important molecular components of centrosomes where discovered in the last 2 decades, but we still do not fully understand how these proteins work together to ensure the spatiotemporal assembly and function of a bipolar spindle. While most of our knowledge about centrosomes was discovered in tissue cell culture systems or model organisms, we have relatively little insight into the functions of centrosomes in the different human cell types that make up tissues and organs. Recent studies indicate that misregulation of centrosome biogenesis and amplification can lead to developmental defects and cancer. In light of these findings, it is important to understand how organisms prevent centrosome alterations to maintain cell and tissue integrity. In this project I proposed to study different aspects of centrosome assembly, function and integrity. To analyze the effect of centrosome removal in normal and cancer cells, I have used centrinone, a potent and specific inhibitor of Polo-like kinase 4 (Plk4), which is essential for centrosome duplication. Across cell types, removing centrosomes by inhibiting Plk4 prolongs mitosis by increasing the time required for spindle assembly. However, cancer cells have a broad range of responses to centrosome removal. While some cancer cell types can divide almost with similar kinetics with and without centrosomes, other cell types have severe problems to progress through mitosis without centrosomes and eventually die. To identify factors that dictate susceptibility to Plk4 inhibition, I studied molecular mechanisms of acentriolar spindle formation. My work suggests that there is an alternative pathway in some cell types that supports the formation of centrosome-like foci containing pericentriolar material (PCM) proteins that can nucleate microtubules and drive spindle assembly when centrosomes are removed. While some cancer cells depend on the ability to form these foci to be able to assemble a spindle when centrosomes are removed, other cells can drive spindle assembly independently of PCM foci. In cells with an intact p53 tumor suppressor pathway, prolonged mitosis leads to p53 stabilization and senescence or cell death. This mechanism is most likely in place to protect cells from accumulating defects that can lead to oncogenic transformation and tumorigenesis. My work on human embryonic stem cells also suggests that asymmetric Wnt signaling can prolong mitosis to allow orientation of the mitotic spindle. In a genome-wide CRISPR/Cas9 loss-of-function screen for centrinone resistance, I identified the first molecular components (53BP1 and USP28) of a pathway that ensures the fidelity of cell division by stabilizing p53 when mitosis is prolonged. This screen also identified the ubiquitin ligase TRIM37, whose loss facilitates mitosis in centrosomeless cells. I showed that loss of TRIM37 potentiates the assembly of the PCM foci that can catalyze spindle assembly. In cells with centrioles, TRIM37 holds PCM assembly in check to ensure that it only forms in the vicinity of centrosomes. TRIM37 overexpression prevents the formation of centrosomes in cells with centrioles and increases mitotic defects by suppressing the formation of PCM foci in cells that lack centrioles. Consistent with these results, cancer cell lines in which TRIM37 is amplified, are highly sensitive to centrosome removal, which causes high levels of mitotic failure. Thus, my results suggest that centrosome removal is a potential strategy for treatment of cancers with high TRIM37 expression.
Publications
- (2016). 53BP1 and USP28 mediate p53 activation and G1 arrest after centrosome loss or extended mitotic duration. Journal of Cell Biology 214, 155-166
Meitinger, F., Anzola, J.V., Kaulich, M., Richardson, A., Stender, J.D., Benner, C., Glass, C.K., Dowdy, S.F., Desai, A., Shiau, A.K., et al.
(See online at https://doi.org/10.1083/jcb.201604081) - (2017). Kinetochores accelerate or delay APC/C activation by directing Cdc20 to opposing fates. Genes & Development 31, 1089-1094
Kim, T., Lara-Gonzalez, P., Prevo, B., Meitinger, F., Cheerambathur, D.K., Oegema, K., and Desai, A.
(See online at https://doi.org/10.1101/gad.302067.117) - (2018). NDF, a Nucleosome Destabilizing Factor that Facilitates Transcription through Nucleosomes. Genes & Development 32, 682–694
Fei, J., Ishii, H., Kassavetis, G.A., Hoeksema, M.A., Meitinger, F., Glass, C.K., Ren, B., and Kadonaga, J.T.
(See online at https://doi.org/10.1101/gad.313973.118)
