Vasquez R J, Gard D L, Cassimeris L
Department of Molecular Biology, Lehigh University, Bethlehem, Pennsylvania 18015.
J Cell Biol. 1994 Nov;127(4):985-93. doi: 10.1083/jcb.127.4.985.
We have used video-enhanced DIC microscopy to examine the effects of XMAP, a Mr 215,000 microtubule-associated protein from Xenopus eggs (Gard, D.L., and M. W. Kirschner. 1987. J. Cell Biol. 105:2203-2215), on the dynamic instability of microtubules nucleated from axoneme fragments in vitro. Our results indicate that XMAP substantially alters the parameters of microtubule assembly at plus ends. Specifically, addition of 0.2 microM XMAP resulted in (a) 7-10-fold increase in elongation velocity, (b) approximately threefold increase in shortening velocity, and (c) near elimination of rescue (the switch from rapid shortening to elongation). Thus, addition of XMAP resulted in the assembly of longer, but more dynamic, microtubules from the plus ends of axonemes which upon catastrophe disassembled back to the axoneme nucleation site. In agreement with previous observations (Gard, D.L., and M. W. Kirschner. 1987. J. Cell Biol. 105:2203-2215), the effects of XMAP on the minus end were much less dramatic, with only a 1.5-3-fold increase in elongation velocity. These results indicate that XMAP, unlike brain MAPs, promotes both polymer assembly and turnover, and suggests that the interaction of XMAP with tubulin and the function of XMAP in vivo may differ from previously characterized MAPs.
我们使用视频增强型微分干涉差显微镜来研究XMAP对体外轴丝片段成核的微管动态不稳定性的影响。XMAP是一种来自非洲爪蟾卵的分子量为215,000的微管相关蛋白(Gard, D.L., and M. W. Kirschner. 1987. J. Cell Biol. 105:2203 - 2215)。我们的结果表明,XMAP显著改变了微管正端的组装参数。具体而言,添加0.2微摩尔的XMAP导致:(a)伸长速度增加7 - 10倍;(b)缩短速度增加约三倍;(c)救援现象(从快速缩短转变为伸长)几乎消失。因此,添加XMAP导致从轴丝正端组装出更长但更具动态性的微管,这些微管在发生灾变时会解体回到轴丝成核位点。与先前的观察结果一致(Gard, D.L., and M. W. Kirschner. 1987. J. Cell Biol. 105:2203 - 2215),XMAP对负端的影响要小得多,伸长速度仅增加1.5 - 3倍。这些结果表明,与脑微管相关蛋白不同,XMAP既能促进聚合物组装又能促进周转,这表明XMAP与微管蛋白的相互作用以及其在体内的功能可能与先前已表征的微管相关蛋白不同。