Microtubule dynamics in mitotic spindles of living cells.
Microtubule dynamics in mitotic spindles of living cells.
复制标题
活细胞有丝分裂纺锤体的微管动力学。
DOI：
10.1111/j.1749-6632.1986.tb38434.x
复制
发表时间：
1986
影响因子：
5.2
通讯作者：
Salmon,ED
中科院分区：
文献类型：
--
作者：
Wadsworth,P;Salmon,ED
《关于活的，分裂的细胞的观察》，以及最近在体外组装的微管(MT)的实验(请参阅参考文献4)，表明MT是动态的结构，对这种动态行为的理解对于理解有丝分裂至关重要。早期对活细胞的研究清楚地证明了含MT的纺锤体纤维和微管蛋白亚基的细胞池之间的不稳定平衡。此外，这种组装和拆解在染色体运动中的作用是这些研究表明，纺锤体MT的组装和随后的拆卸与有丝分裂过程密切相关。然而，微管蛋白与纺锤体MTS结合和解离的途径和调节尚未解决。使用免疫荧光和电子显微镜进行的各种形态研究表明，纺锤体部分由从纺锤体极辐射并在中期板重叠的MTS的两极排列组成。然而，这种技术只能产生固定染色细胞的静态图像，因此MTS对染色体运动的贡献仍然不清楚。为了研究纺锤体MT的动力学，我们开发了将荧光标记的微管蛋白亚单位引入活的技术，分裂细胞以监测微管蛋白的行为。这项工作是与J·R·麦金托什的实验室合作进行的(本卷第566页)。使用这种被称为荧光模拟细胞化学的方法，可以研究活细胞中微管蛋白亚单位的分布和动态，避免了复杂的体外条件。14岁的Keith et A1最初使用了类似的方法，他们观察了培养细胞中间期MT阵列的动态行为。随后，Wadsworth和Sloboda“展示了在有丝分裂过程中，海胆卵中的荧光微管蛋白快速聚合成MT阵列。这些初步研究表明，这些技术可用于检测活细胞中微管蛋白的行为。在我们的实验中，牛脑微管蛋白已经被5-(4，6-二氯三氮唑-2-基)氨基荧光素(DTAF)共价修饰，得到的DTAF微管蛋白被证明保留了微管蛋白分子的固有特征。(标记过程的细节可以在参考文献16和J.R.McIntosh，这卷566页中找到)。在显微注射到合适的细胞后，DTAF-微管蛋白已被证明与活细胞中所有含有MT的细胞质结构共同组装。18因此，外源标记的亚基可作为内源性微管蛋白池行为的“示踪剂”。
Observations on living, dividing cells,’” together with more recent experiments with microtubules (MTs) assembled in vitro (for review see reference 4), suggest that MTs are dynamic structures and that an understanding of this dynamic behavior is critical for an understanding of mitosis. Early work on living cells clearly demonstrated the labile equilibrium between the MT-containing spindle fibers and a cellular pool of tubulin subunits. In addition, a role for this assembly and disassembly in chromosome movement was These studies showed that the assembly of spindle MTs and their subsequent disassembly are intimately linked to the mitotic process. The pathways and regulation of tubulin association and dissociation with spindle MTs, however, were not resolved. A variety of morphological studies, performed using immunofluorescence and electron microscopy, have revealed that spindles are composed, in part, of a bipolar arrangement of MTs that radiate from the spindle poles and overlap at the metaphase plate. 612 Such techniques, however, yield only static images of fixed stained cells, so the contribution of MTs to chromosome movement has remained unclear.In an effort to investigate spindle MT dynamics, we have developed techniques to introduce fluorochrome-labeled tubulin subunits into living, dividing cells to monitor tubulin behavior. This work has been carried out in collaboration with the laboratory of J. R. McIntosh (this volume p. 566). Using this approach, termed fluorescence analogue cytochemistry,” the distribution and dynamics of the tubulin subunit can be investigated in living cells, avoiding the complexities of in vitro conditions. Similar methods were initially used by Keith et a1., 14 who observed the dynamic behavior of the interphase MT array in cultured cells. Subsequently, Wadsworth and Sloboda” demonstrated the rapid polymerization of fluorescent tubulin into MT arrays in sea urchin eggs during mitosis. These preliminary studies suggested the utility of these techniques to examine the behavior of tubulin in living cells. For our experiments, bovine brain tubulin has been covalently modified with the fluorochrome 5-(4, 6-dichlorotriazin-2-yl) amino fluorescein (DTAF) and the resulting DTAF tubulin shown to retain the native characteristics of the tubulin molecule.(Details of the labeling procedures can be found in reference 16 and J. R. McIntosh, this volume p. 566). Following microinjection into suitable cells, the DTAF-tubulin has been shown to coassemble with all cytoplasmic MT containing structures in living cells.”. 18 Thus the exogenous labeled subunits serve as “tracers” of the behavior of the endogenous tubulin pool.
登录
查看更多内容
影响因子：
3.9
作者：
HEPLER, PK;CLEARY, AL;ZHANG, DH
通讯作者：
ZHANG, DH
影响因子：
64.5
作者：
LAMB, NJC;FERNANDEZ, A;CAVADORE, JC
通讯作者：
CAVADORE, JC
植物细胞中微管蛋白和微管阵列的免疫荧光显微镜检查。
DOI：
--
发表时间：
1985
期刊：
Cell Biology International Reports
影响因子：
--
作者：
Susan M. Wick
通讯作者：
Susan M. Wick
高等植物中的细胞分裂：豌豆中的 cdc2 基因、其 34-kDa 产物和组蛋白 H1 激酶活性。
DOI：
10.1073/pnas.87.14.5397
发表时间：
1990
影响因子：
11.1
作者：
H. Feiler;Thomas Jacobs
通讯作者：
Thomas Jacobs
微管蛋白荧光类似物的制备和表征。
影响因子：
--
作者：
P. Wadsworth;E. Salmon
通讯作者：
E. Salmon