CENTROMERE-DEPENDENT BINDING OF YEAST MINICHROMOSOMES TO MICROTUBULES INVITRO
CENTROMERE-DEPENDENT BINDING OF YEAST MINICHROMOSOMES TO MICROTUBULES INVITRO
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DOI：
10.1016/0092-8674(81)90012-x
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发表时间：
1991-08-09
期刊：
影响因子：
64.5
通讯作者：
KOSHLAND, D
中科院分区：
文献类型：
--
作者：
KINGSBURY, J;KOSHLAND, D
我们提出了酵母着丝粒功能的体外测定；分离的酵母小染色体需要一个功能的着丝粒来结合牛微管和沉积物。着丝粒-牛微管复合物在生理微管浓度下形成。三个着丝粒DNA元素中的两个，在体内是着丝粒功能所必需的，在体外也是着丝粒结合微管所必需的。然而，纯化的着丝粒DNA单独不与微管结合。这些结果表明，微管结合必须由两个着丝粒DNA元件和与其中一个或两个相关的因子介导。由nocodazolg捕获的G2-M细胞的裂解物中具有微管结合活性的着丝粒的百分比比α因子G1细胞高7- 10倍，这表明这种着丝粒活性在细胞周期中受到调节。讨论了该分析在解剖着丝粒组装、功能和调节方面的潜力。
We present an in vitro assay for yeast centromere function; isolated yeast minichromosomes require a functional centromere to bind to bovine microtubules and sediment with them. Centromere-bovine microtubule complexes form at physiological microtubule concentrations. Two of the three centromere DNA elements, which are necessary for centromere function in vivo, are also necessary for centromeres to bind microtubules in vitro. However, purified centromere DNA alone does not bind to microtubules. These results suggest that microtubule binding must be mediated by the two centromere DNA elements and factors that associate with one or both of them. The percent of centromeres with microtubule-binding activity is 7- to 10-fold higher in lysates made from nocodazole-arrested G2-M cells than from alpha-factor G1 cells, suggesting that this centromere activity is regulated during the cell cycle. The potential of this assay for dissecting centromere assembly, function, and regulation is discussed.