MICROTUBULES OF MITOTIC APPARATUS

本研究的主要目的是：1）鉴定 与有丝分裂器微管直接相关的复合物， 紫球海胆卵：2）调查是否相似 组织存在于有丝分裂器微管结构中， 研究精子鞭毛轴丝的功能和组装 与纺锤体纤维微管相连的亚结构， 星光 该研究将对离散数量的蛋白质进行， 从有丝分裂器制备物中溶解。 那些组件 在特定条件下与微管蛋白一起沿着提取 用于微管解聚。 通过以下方法制备的级分的组分 不同的物理或化学处理将通过一个比较， 双向凝胶电泳 推定的微管相关 多肽应以恒定的摩尔比存在于所有级分中 相对于微管蛋白。 比较分析将扩大到 整个轴丝或多肽优先溶解于不同的 轴丝亚结构 大量的杂交瘤分泌单克隆 针对有丝分裂器多肽和有丝分裂器多肽的抗体， 轴丝，将通过灵敏的固相放射免疫测定法选择。 与 单克隆抗体寻找两种肽之间的相似性， 具有相似的抗原决定簇，可以通过肽分析和 与来自相同分子的其它分子形成的复合物的结构分析 源头 有丝分裂器多肽似乎与 微管将在有丝分裂的不同阶段定位于细胞中， 抗体。 体外结合的抗原的定量分析 将通过放射免疫测定法进行微管。

The major objectives of this project are: 1) to identify the molecular complexes which directly associate with mitotic apparatus microtubules in Strongylocentrotus purpuratus eggs: 2) to investigate whether similar organizations exist in the mitotic apparatus microtubular structures and in the axoneme of sperm flagellum; 3) to study the function and the assembly of substructures connected with the microtubules of the spindle fibers and of the astral rays. The study will be performed on a discrete number of proteins which are solubilized from mitotic apparatus preparations. Those components are extracted along with microtubular proteins under conditions which are specific for microtubule depolymerization. Constituents of fractions prepared by different physical or chemical treatments will be compared by one- and two-dimensional gel electrophoresis. Putative microtubule-associated polypeptides should be present in all the fractions with a constant molar ratio versus the tubulin. The comparative analysis will be extended to components of the whole axoneme or to polypeptides preferentially solubilized from different axonemal substructures. A large number of hybridomas secreting monoclonal antibodies directed against polypeptides of the mitotic apparatus and of the axoneme, will be selected by a sensitive solid phase radioimmunoassay. With the monoclonal antibodies the search for similarities between the two peptides which have similar antigenic determinants could be studied by peptide analysis and by structural analysis of the complexes formed with other molecules from the same source. Mitotic apparatus polypeptides which appear to be associated with microtubules will be localized in the cell during different phases of mitosis by the antibodies. Quantitative analysis of antigens which bind in vitro to microtubules will be performed by radioimmunoassay.