Arquint, Christian. On the function and regulation of human STIL - a centrosomal protein implicated in autosomal recessive primary microcephaly. 2014, PhD Thesis, University of Basel, Faculty of Science.
Official URL: http://edoc.unibas.ch/diss/DissB_11094
Drosophila Ana2 and C. elegans SAS-5 family of centriole duplication factors and a protein, which causes autosomal recessive primary microcephaly (MCPH) when mutated in patients. We show that depletion of STIL from human cells blocks centriole duplication, whereas overexpression of STIL triggers the near-simultaneous formation of multiple daughter centrioles. A similar phenotype had previously been observed for HsSAS-6 and the kinase Plk4, two key regulators of centriole duplication that contribute to cartwheel assembly, a template for centriole formation. In line with these results, we observed a prominent co-localization of STIL and HsSAS-6 at the cartwheel region. Together with two independent studies (Tang et al., 2011; Vulprecht et al., 2012), our work suggests that STIL cooperates with Plk4 and HsSAS-6 in cartwheel formation and thus represents a key centriole duplication factor in human cells.
The observation that excess STIL triggers centriole amplification, a condition that is associated with genome instability, prompted us to analyse the controls governing STIL cell-cycle regulation in more detail. By fluorescence time-lapse imaging, we revealed a two-step process that results in complete elimination of STIL towards the end of mitosis. First, during nuclear envelope breakdown, Cdk1 triggers the translocation of STIL from daughter centrioles into the cytoplasm. This event might initiate cartwheel disassembly, as HsSAS-6, a major cartwheel component, follows a similar trend. The bulk of cytoplasmic STIL is then degraded at the metaphase to anaphase transition by the anaphase promoting complex/cyclosome (APC/C), which involves a KEN box located at the C-terminus of STIL. Interestingly, we found that truncations of STIL that cause MCPH in human patients delete this KEN box, but preserve the overall function of STIL as a centriole duplication factor. We readily confirmed that STIL MCPH truncations resist APC/C-mediated degradation, and demonstrated that stabilization of mutant STIL is strong enough to trigger centriole amplification in our cell culture model. Therefore, by analysing STIL cell cycle regulation, we uncovered a provocative link to primary microcephaly. This leads us to propose that centriole amplification, triggered by STIL stabilization, is the underlying cause of MCPH in patients with STIL mutations.
|Advisors:||Nigg, Erich A.|
|Committee Members:||Affolter, Markus|
|Faculties and Departments:||05 Faculty of Science > Departement Biozentrum > Growth & Development > Cell Biology (Nigg)|
|Bibsysno:||Link to catalogue|
|Number of Pages:||140 S.|
|Last Modified:||30 Jun 2016 10:56|
|Deposited On:||28 Jan 2015 15:32|
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