We have isolated chromosomes from Chinese hamster ovary cells arrested in mitosis with vinblastine and examined the interactions of their kinetochores with purified tubulin in vitro. The kinetochores nucleate microtubule (MT) growth with complex kinetics. After an initial lag phase, MTs are continuously nucleated with both plus and minus ends distally localized. This mixed polarity seems inconsistent with the formation of an ordered, homopolar kinetochore fiber in vivo. As isolated from vinblastine-arrested cells, kinetochores contain no bound tubulin. The kinetochores of chromosomes isolated from colcemid-arrested cells or of chromosomes incubated with tubulin in vitro are brightly stained after anti-tubulin immunofluorescence. This bound tubulin is probably not in the form of MTs. It is localized to the corona region by immunoelectron microscopy, where it may play a role in MT nucleation in vitro.
Article| September 01 1985
Properties of the kinetochore in vitro. I. Microtubule nucleation and tubulin binding.
T J Mitchison
M W Kirschner
Online ISSN: 1540-8140
Print ISSN: 0021-9525
J Cell Biol (1985) 101 (3): 755–765.
T J Mitchison, M W Kirschner; Properties of the kinetochore in vitro. I. Microtubule nucleation and tubulin binding.. J Cell Biol 1 September 1985; 101 (3): 755–765. doi: https://doi.org/10.1083/jcb.101.3.755
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