122 research outputs found
PLK1 controls centriole distal appendage formation and centrobin removal via independent pathways
No full textPrimary and Secondary data linked to the publication "PLK1 controls centriole distal appendage formation and centrobin removal via independent pathways" in Journal of Cell Science, Le Roux-Bourdieu et al., 202
Mild replication stress causes premature centriole disengagement via a sub-critical Plk1 activity under the control of ATR-Chk1
No full textDataset for the paper Mild replication stress causes premature centriole disengagement via a sub-critical Plk1 activity under the control of ATR-Chk
Centrosome age breaks spindle size symmetry even in “symmetrically” dividing cells
No full textOpen access data for the publication "Centrosome age breaks spindle size symmetry even in “symmetrically” dividing cells" in the Journal of Cell Biolog
Evidence for a HURP/EB free mixed-nucleotide zone in kinetochore-microtubules
No full textDataset of the publication "Evidence 1 for a HURP/EB free mixed-nucleotide zone in kinetochore-microtubules" by Castrogiovanni, Inchingolo et al in Nature Communicaiton 202
Control of chromosome biorientation by a feedback loop involving the Ska complex and Aurora B kinase
No full textProper chromosome attachment to opposite spindle poles (biorientation) and error-free chromosome segregation relies on the plasticity of kinetochore-microtubule attachments; these must remain flexible enough to allow the release of erroneously attached spindle microtubules, yet become sufficiently stable to harness forces for chromosome movements and silence the spindle assembly checkpoint. Aurora B kinase fosters chromosome biorientation by facilitating the dynamics of kinetochore-microtubule attachments through phosphorylation of kinetochore proteins that bind microtubules. Prominent among the substrates, whose microtubule and kinetochore binding is curtailed by Aurora B, is the Ska complex, a key factor for kinetochore-fiber stability. Here, we show that Ska is not only a substrate of Aurora B, but is also required, in turn, for Aurora B activity. Ska-deficient cells fail to biorient and display lagging chromosomes and micronuclei as a result of suppressed kinetochore-microtubule turnover. These defects coincide with diminished kinetochore localization of the Aurora B effectors MCAK and CENP-E, as well as reduced Aurora B substrate phosphorylation. We further show that Ska requires its microtubule binding capability to promote Aurora B activity in cells and directly stimulates Aurora B catalytic activity in vitro. Finally, we demonstrate that PP1 counters Aurora B activity to enable Ska kinetochore accumulation once biorientation is achieved, which allows Ska to exert its kinetochore-fiber stabilizing function. Together, we propose that the Ska complex enhances Aurora B activity to limit its own microtubule and kinetochore association and ensure that the dynamics and stability of kinetochore-microtubules fall within an optimal balance for chromosome biorientation and faithful chromosome segregation
Phylogenetic and structural analysis of centromeric DNA and kinetochore proteins
No full textBackground: Kinetochores are large multi-protein structures that assemble on centromeric DNA (CEN DNA) and mediate the binding of chromosomes to microtubules. Comprising 125 base-pairs of CEN DNA and 70 or more protein components, Saccharomyces cerevisiae kinetochores are among the best understood. In contrast, most fungal, plant and animal cells assemble kinetochores on CENs that are longer and more complex, raising the question of whether kinetochore architecture has been conserved through evolution, despite considerable divergence in CEN sequence. Results: Using computational approaches, ranging from sequence similarity searches to hidden Markov model-based modeling, we show that organisms with CENs resembling those in S. cerevisiae (point CENs) are very closely related and that all contain a set of 11 kinetochore proteins not found in organisms with complex CENs. Conversely, organisms with complex CENs (regional CENs) contain proteins seemingly absent from point-CEN organisms. However, at least three quarters of known kinetochore proteins are present in all fungi regardless of CEN organization. At least six of these proteins have previously unidentified human orthologs. When fungi and metazoa are compared, almost all have kinetochores constructed around Spc105 and three conserved multi-protein linker complexes (MIND, COMA, and the NDC80 complex). Conclusion: Our data suggest that critical structural features of kinetochores have been well conserved from yeast to man. Surprisingly, phylogenetic analysis reveals that human kinetochore proteins are as similar in sequence to their yeast counterparts as to presumptive Drosophila melanogaster or Caenorhabditis elegans orthologs. This finding is consistent with evidence that kinetochore proteins have evolved very rapidly relative to components of other complex cellular structures.Jane Coffin Childs Fund for Medical ResearchEuropean Molecular Biology OrganizationNational Institutes of Health (U.S.) (Grant CA84179) (Grant GM51464
Regulation of chromosome congression : focus on the function of hSpindly and the kinetochore recruitment of the Ska complex
No full textMitosis is a process in which cells divide their genetic materials equally into two daughter cells. During mitosis, specialized structures called kinetochores (KTs), located at the centromeric regions of chromosomes, are captured by microtubules (MTs) radiated from spindle poles. Subsequently, the chromosomes align at the metaphase plate, a process called congression. This process needs to be tightly controlled in order to maintain genomic stability, as subsequent chromosome segregation depends critically on KT-MT interactions. However, how KT-MT attachments are regulated remains largely unknown. Therefore, identification of novel KT/spindle proteins and thorough examination of their functions and regulation will increase our understanding on chromosome congression mechanisms and thus mitotic progression.
This thesis focuses on the study of two KT and spindle components previously identified in a survey of the human mitotic spindle and it is therefore divided into two parts. The first part describes the functional characterization of a novel KT and spindle localizing protein, human Spindly (hSpindly, previously called CCDC99), which is the human homologue of Drosophila Spindly. We show that hSpindly specifically recruits dynein/dynactin to KTs. Localization of hSpindly is in turn controlled by the Rod/ZW10/Zwilch (RZZ) complex and Aurora B kinase. hSpindly depletion results in reduced inter-KT tension, unstable KT-MT fibers (K-fibers), and extensive prometaphase delay and severe chromosome misalignment. Moreover, depletion of hSpindly induces a striking spindle rotation, which can be rescued by co-depletion of dynein. However, in contrast to Drosophila, hSpindly depletion does not abolish the removal of MAD2 and ZW10 from KTs. Collectively, our data reveal hSpindly-mediated dynein functions and highlight a critical role of KT dynein in spindly orientation.
In the second part of this thesis, the regulation of the Ska complex (composed of Ska1, Ska2 and Ska3) has been studied. We show that Aurora B activity negatively regulates the localization of the Ska complex to KTs. Furthermore, recruitment of the Ska complex to KTs depends on the KNL-1/Mis12/Ndc80 (KMN) network. In agreement with this, we have identified interactions between members of the KMN and Ska complexes and demonstrate that the interaction between the two complexes is regulated by Aurora B activity. Aurora B can directly phosphorylate Ska1 and Ska3 in vitro, and expression of phosphomimetic mutants of Ska1 and Ska3 impairs Ska KT recruitment and formation of stable K-fibers, disrupting mitotic progression. We propose that Aurora B phosphorylation antagonizes the interaction between the Ska complex and the KMN network, thereby controlling Ska KT recruitment and stabilization of KT-MT attachments.
Together, we conclude that hSpindly and the Ska complex, two important components of KTs in metazoans, are involved in the regulation of chromosome congression by recruiting KT dynein and stabilizing KT-MT attachments, respectively. Both their function and localization are tightly regulated by mitotic kinases and upstream structural components
Keeping kinetochores on track
No full textThe multiple functions of kinetochores are reflected in their complex composition, with over a hundred different proteins, which self-associate in several functional subcomplexes. Most of these kinetochore proteins were identified over the last 10-12 years using a combination of genetic, cell biological, biochemical, and bioinformatic approaches in various model organisms. The key challenge since then has been to determine the structural architecture of kinetochores, define the functions of its different subcomponents, and understand its regulation, both in response to the rapid changes in microtubule dynamics or to sense erroneous attachments for spindle checkpoint signalling. Here, we present some of the key advances obtained in the last six years on the biology of kinetochores, both through our work and through the work of many other groups studying this exciting structure
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