Interzone microtubule behavior in late anaphase and telophase spindles.
Interzone microtubule behavior in late anaphase and telophase spindles.
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DOI：
10.1083/jcb.105.2.875
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发表时间：
1987-08
期刊：
影响因子：
--
通讯作者：
McIntosh JR
中科院分区：
文献类型：
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作者：
Saxton WM;McIntosh JR
我们研究了微管行为后期和末期纺锤体的PtK 1细胞，使用荧光微管蛋白（DTAF-tubulin），显微注射，和激光微束光漂白。我们提出了两个新的测试结果，增加了证据表明，DTAF-微管蛋白密切模仿本地微管蛋白在体内的行为。(a)显微注射DTAF微管蛋白与注射天然微管蛋白在促进被剥夺紫杉醇的紫杉醇依赖性有丝分裂突变细胞分裂方面一样有效。(b)显微注射秋水仙素-DTAF-微管蛋白复合物与注射秋水仙素-天然微管蛋白复合物在引起纺锤体解聚方面相似。在后期或末期将DTAF-微管蛋白显微注射到野生型细胞中后，立即在染色体半纺锤体和染色体后面观察到微管的荧光掺入，但在中间区的中间附近没有荧光掺入。在接下来的几分钟里，微管蛋白荧光在中间区（赤道）的中心聚集，变得越来越强烈。在其他实验中，细胞显微注射与DTAF-微管蛋白在前期，并允许平衡30分钟。细胞已进展到后期，然后进行光漂白，以减少在中间区的中心部分的荧光。在几分钟的时间内，荧光的唯一实质性重新分布是在中间区的赤道处出现明亮区域。荧光掺入和光漂白数据的网站表明，微管蛋白添加到赤道附近的细长纺锤体中间区，其中的交叉指状微管的正端位于。在进一步的实验中，几条暗线垂直于荧光后期-末期纺锤体的极-极轴被光漂白。随时间变化的线之间的间距表明，两个半的中间地带躺在相对两侧的纺锤体赤道移动远离彼此。这表明，构成大部分中间区的交叉指状微管可以进行反平行滑动。我们的数据支持后期B的模型，其中加上末端延伸的交叉微管和反平行滑动有助于染色体分离。
We have studied microtubule behavior in late anaphase and telophase spindles of PtK1 cells, using fluoresceinated tubulin (DTAF-tubulin), microinjection, and laser microbeam photobleaching. We present the results of two novel tests which add to the evidence that DTAF-tubulin closely mimics the behavior of native tubulin in vivo. (a) Microinjected DTAF-tubulin was as effective as injected native tubulin in promoting division of taxol-dependent mitotic mutant cells that had been deprived of taxol. (b) Microinjected colchicine-DTAF-tubulin complex was similar to injected colchicine-native tubulin complex in causing depolymerization of spindles. Immediately after microinjection of DTAF-tubulin into wild-type cells during late anaphase or telophase, fluorescence incorporation by microtubules was seen in chromosomal half- spindles and just behind the chromosomes, but there was no fluorescence incorporation near the middle of the interzone. Over the next few minutes, tubulin fluorescence accumulated at the center of the interzone (the equator), becoming progressively more intense. In other experiments, cells were microinjected with DTAF-tubulin at prophase and allowed to equilibrate for 30 min. Cells that had progressed to late anaphase were then photobleached to reduce the fluorescence in the central portion of the interzone. Over a period of several minutes, the only substantial redistribution of fluorescence was the appearance of a bright area at the equator of the interzone. Both the site of fluorescence incorporation and the photobleaching data suggest that tubulin adds to the elongating spindle interzone near the equator where the plus ends of the interdigitated microtubules are located. In further experiments, several dark lines were photobleached perpendicular to the pole-to-pole axis of fluorescent anaphase- telophase spindles. Time-dependent changes in the spacings between the lines indicated that the two halves of the interzone lying on opposite sides of the spindle equator moved away from one another. This shows that the interdigitated microtubules, which make up most of the interzone, can undergo antiparallel sliding. Our data support a model for anaphase B in which plus end elongation of interdigitated microtubules and antiparallel sliding contribute to chromosome separation.