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- W830367544 abstract "The mitotic spindle is required to accurately separate the two duplicated copies of the genome between the two daughter cells. Errors that occur during this process can lead to aneuploidy, a hallmark of cancer. Some microtubules of the spindle are bundled by inter-microtubule bridges to form kinetochore fibres (K-fibres), which are responsible for pulling the sister chromatids apart. Previous work has identified one type of inter- microtubule bridge, composed of three proteins: TACC3, ch-TOG and clathrin. These TACC3–ch-TOG–clathrin crosslinkers are important for accurate mitosis and were found to contribute to K-fibre stability.However, these studies used RNAi-mediated depletion to study the functions of these proteins at metaphase. RNAi-mediated depletion is slow, and it is therefore difficult to separate the roles of the proteins at early mitosis versus metaphase. For instance, the depletion phenotype at metaphase may have been due to the absence of the protein at metaphase, or to the assembly of a defective mitotic spindle in early mitosis.Here, knocksideways (KS) was used to deplete TACC3 in under 5 minutes from fully assembled spindle at metaphase, and trap the protein on the external membrane of mitochondria. TACC3 KS also caused the rerouting of both ch-TOG and clathrin, without affecting other spindle proteins. This depletion was supported by a concomitant reduction of inter-microtubule bridge frequency in K-fibres. When TACC3 KS was performed at early mitosis, cells displayed severe delays in aligning chromosomes. When TACC3 KS was performed at metaphase following a normal prometaphase, anaphase onset was delayed. In these cells, Mad2 on a subset of kinetochores indicated that the spindle checkpoint was active, and K-fibre tension was reduced. However, both a cold-stable K-fibre assay and electron microscopy analysis of K-fibres indicated that there was no significant loss of K-fibre microtubules, even after prolonged removal at metaphase. Furthermore, TACC3 KS at metaphase significantly altered kinetochore microdynamics and reduced spindle length.These results indicate that TACC3–ch-TOG–clathrin crosslinkers are important for mitotic spindle assembly, but not for K-fibre stability at metaphase. However, the complex is required for K-fibre function at metaphase, satisfaction of the spindle checkpoint, most likely by maintenance of K-fibre tension." @default.
- W830367544 created "2016-06-24" @default.
- W830367544 creator A5004911952 @default.
- W830367544 date "2013-09-01" @default.
- W830367544 modified "2023-09-26" @default.
- W830367544 title "Rapid removal of TACC3–ch-TOG–clathrin from kinetochore fibres during mitosis" @default.
- W830367544 hasPublicationYear "2013" @default.
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