The Kinetochore-Bound Ska1 Complex Tracks Depolymerizing Microtubules and Binds to Curved Protofilaments

Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
Developmental Cell (Impact Factor: 9.71). 10/2012; 23(5). DOI: 10.1016/j.devcel.2012.09.012
Source: PubMed

ABSTRACT To ensure equal chromosome segregation during mitosis, the macromolecular kinetochore must remain attached to depolymerizing microtubules, which drive chromosome movements. How kinetochores associate with depolymerizing microtubules, which undergo dramatic structural changes forming curved protofilaments, has yet to be defined in vertebrates. Here, we demonstrate that the conserved kinetochore-localized Ska1 complex tracks with depolymerizing microtubule ends and associates with both the microtubule lattice and curved protofilaments. In contrast, the Ndc80 complex, a central player in the kinetochore-microtubule interface, binds only to the straight microtubule lattice and lacks tracking activity. We demonstrate that the Ska1 complex imparts its tracking capability to the Ndc80 complex. Finally, we present a structure of the Ska1 microtubule-binding domain that reveals its interaction with microtubules and its regulation by Aurora B. This work defines an integrated kinetochore-microtubule interface formed by the Ska1 and Ndc80 complexes that associates with depolymerizing microtubules, potentially by interacting with curved microtubule protofilaments.

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Available from: Ekaterina L Grishchuk, Jan 04, 2014
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    • "Such a mechanism must involve proteins that (1) mediate end-on microtubule–kinetochore attachments, (2) maintain attachment to depolymerising microtubules and (3) ensure the K-fibre remains in a net depolymerising state through control of kinetochore–microtuble dynamics. The Ndc80 and Ska complexes can directly bind to microtubules in vitro, with the Ska complex (but not the Ndc80 complex) able to autonomously track depolymerising microtubule plus-ends (Schmidt et al., 2012). Extensive depletion of either complex results in a severe failure in chromosome congression and the inability to form a metaphase plate (Daum et al., 2009; Gaitanos et al., 2009; Welburn et al., 2010). "
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    ABSTRACT: A key step of mitosis is the congression of chromosomes to the spindle equator. Congression is driven by at least two distinct mechanisms: (1) kinetochores slide along the microtubule lattice using the plus-end directed CENP-E motor, and (2) kinetochores biorientating near the pole move to the equator through microtubule depolymerisation-coupled pulling. Here, we show that CENP-Q - a subunit of the CENP-O/P/Q/U complex that targets polo-like kinase (Plk1) to kinetochores - is also required for the recruitment of CENP-E to kinetochores. We further reveal a CENP-E recruitment-independent role for CENP-Q in depolymerisation-coupled pulling. Both these functions are abolished by a single point mutation in CENP-Q (S50A) - a residue that is phosphorylated in vivo. Importantly, the S50A mutant does not affect Plk1 loading onto kinetochores and leaves the CENP-O complex intact. Thus, the function of CENP-Q in CENP-E loading and depolymerisation-coupled pulling are independent from its role in Plk1 recruitment and CENP-O/P/Q/U complex stabilization. Together, our data provide evidence that phospho-regulation of CENP-Q plays a central function in coordinating chromosome congression mechanisms.
    Journal of Cell Science 11/2014; 128(1). DOI:10.1242/jcs.163659 · 5.43 Impact Factor
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    • "Overexpression of AURK-A has been shown to be associated with aneuploidy, chromosome instablity, tumorigenic transformation and progression in mammalian cells (Bischoff et al. 1998; revised in Gómez-López et al. 2014). AURK-B has been shown to regulate SKA1 (Schmidt et al. 2012). KIF11 is a protein required from prophase until metaphase which participates in spindle assembly, centrosome separation, post-mitotic centrosome movement and bipolar spindle formation. "
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    ABSTRACT: Approximately 15% of eukaryotes contain supernumerary B chromosomes. When present, B chromosomes frequently represent as much as 5% of the genome. Despite thousands of reports describing the distribution of supernumeraries in various taxa, a comprehensive theory for the origin, maintenance and evolution of B chromosomes has not emerged. Here we sequence the complete genomes of individual cichlid fish (Astatotilapia latifasciata) with and without B chromosomes, as well as microdissected B chromosomes, to identify DNA sequences on the B. B sequences were further analyzed through qPCR and in situ hybridization. We find that the B chromosome contains thousands of sequences duplicated from essentially every chromosome in the ancestral karyotype. Although most genes on the B chromosome are fragmented, a few are largely intact and we detect evidence that at least three of them are transcriptionally active. We propose a model in which the B chromosome originated early in the evolutionary history of Lake Victoria cichlids from a small fragment of one autosome. DNA sequences originating from several autosomes, including protein-coding genes and transposable elements, subsequently inserted into this proto-B. We propose that intact B chromosome genes involved with spindle fusion and kinetochore attachment may play a role in driving the transmission of the B chromosome. Furthermore, our work suggests that karyotyping is an essential step prior to genome sequencing to avoid problems in genome assembly as well as analytical biases created by the presence of high copy number sequences on the B chromosome.
    Molecular Biology and Evolution 04/2014; 31(8):2061-2072. DOI:10.1093/molbev/msu148 · 9.11 Impact Factor
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    • "Notably, spindle and kinetochore-associated protein complexes have several key properties, and play an important role in coupling chromosome movement to microtubule depolymerization [11,12]. Spindle and kinetochore associated complex subunit 1 (SKA1) is a microtubule-binding subcomplex of the outer kinetochore that is essential for proper chromosome segregation [13,14]. Depletion of SKA1 proteins results in sparse microtubule arrays that often exhibited twisted or bent spindles. "
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    ABSTRACT: The prognosis of oral squamous cell carcinoma is very poor due to local recurrence and metastasis. This study explores the molecular events involved in oral carcinoma with the goal of developing novel therapeutic strategies. The mitotic spindle is a complex mechanical apparatus required for the accurate segregation of sister chromosomes during mitosis. Spindle and kinetochore associated complex subunit 1 (SKA1) is a microtubule-binding subcomplex of the outer kinetochore that is essential for proper chromosome segregation. In recent years, much attention has been focused on determining how SKA proteins interact with each other, as well as their biological role in cancer cells. However, the precise role of SKA1 in oral carcinoma remains unknown. In order to investigate the role of SKA1 in oral cancer, we employed lentivirus-mediated shRNA to silence SKA1 expression in the CAL-27 human oral adenosquamous carcinoma cell line. Depletion of SKA1 in CAL-27 cells significantly decreased cell proliferation, as determined by MTT and colony formation assays. These results strongly demonstrate that reduced SKA1 protein levels may cause inhibition of tumor formation. The shRNA-mediated depletion of SKA1 also led to G2/M phase cell cycle arrest and apoptosis. This is the first report to show that SKA1 plays an important role in the progression of oral adenosqamous carcinoma. Thus, silencing of SKA1 by RNAi might be a potential therapy for this disease.
    Cancer Cell International 08/2013; 13(1):83. DOI:10.1186/1475-2867-13-83 · 2.77 Impact Factor
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