A monoclonal antibody to a mitotic microtubule-associated protein blocks mitotic progression.
A monoclonal antibody to a mitotic microtubule-associated protein blocks mitotic progression.
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DOI：
10.1083/jcb.111.2.511
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发表时间：
1990-08
期刊：
影响因子：
--
通讯作者：
McIntosh JR
中科院分区：
文献类型：
--
作者：
Nislow C;Sellitto C;Kuriyama R;McIntosh JR
从CHO细胞中分离出抗有丝分裂纺锤体的单克隆抗体[CHO1]， Sellitto， C.和R. Kuriyama. 1988。J.细胞生物学，106:431-439)鉴定了一个位于95和105 kD多肽上的表位，并定位于多种生物的纺锤体中。抗原在中期分布于整个纺锤体，但随着后期的进展，在纺锤体中间面两侧的一个逐渐狭窄的区域集中。微量注射CHO1，无论是作为腹水还是作为纯化的IgM，都以阶段特异性和剂量依赖性的方式导致有丝分裂抑制。平行对照注射非免疫igm不产生显著的有丝分裂抑制。注射细胞的免疫荧光分析显示，完成有丝分裂的细胞显示CHO1的正常定位，而阻滞的细胞在纺锤体内没有CHO1抗原的特异性定位。这些细胞的免疫电镜图像显示异常的微管组织。HeLa细胞提取物中的CHO1抗原与紫杉醇稳定的微管相结合。这两种携带抗原的多肽都不能通过ATP或GTP从微管中提取，但在0.5 M NaCl溶液中提取率均约为60%。蔗糖梯度分析表明抗原在11S左右沉积。CHO 1抗原似乎是一种新的有丝分裂MAP，其在纺锤体内的适当分布是有丝分裂所必需的。抗原的特性表明，相应的蛋白质是在后期将两个交叉的半纺锤体的反平行微管保持在一起的机制的一部分。
A monoclonal antibody raised against mitotic spindles isolated from CHO cells ([CHO1], Sellitto, C., and R. Kuriyama. 1988. J. Cell Biol. 106:431-439) identifies an epitope that resides on polypeptides of 95 and 105 kD and is localized in the spindles of diverse organisms. The antigen is distributed throughout the spindle at metaphase but becomes concentrated in a progressively narrower zone on either side of the spindle midplane as anaphase progresses. Microinjection of CHO1, either as an ascites fluid or as purified IgM, results in mitotic inhibition in a stage-specific and dose-dependent manner. Parallel control injections with nonimmune IgMs do not yield significant mitotic inhibition. Immunofluorescence analysis of injected cells reveals that those which complete mitosis display normal localization of CHO1, whereas arrested cells show no specific localization of the CHO1 antigen within the spindle. Immunoelectron microscopic images of such arrested cells indicate aberrant microtubule organization. The CHO1 antigen in HeLa cell extracts copurifies with taxol-stabilized microtubules. Neither of the polypeptides bearing the antigen is extracted from microtubules by ATP or GTP, but both are approximately 60% extracted with 0.5 M NaCl. Sucrose gradient analysis reveals that the antigens sediment at approximately 11S. The CHO 1 antigen appears to be a novel mitotic MAP whose proper distribution within the spindle is required for mitosis. The properties of the antigen(s) suggest that the corresponding protein(s) are part of the mechanism that holds the antiparallel microtubules of the two interdigitating half spindles together during anaphase.