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首页> 外文期刊>Cell motility and the cytoskeleton >Identification and cell cycle-dependent localization of nine novel, genuine centrosomal components in Dictyostelium discoideum.
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Identification and cell cycle-dependent localization of nine novel, genuine centrosomal components in Dictyostelium discoideum.

机译：盘基网柄菌中九种新颖的真正中心体成分的鉴定和细胞周期依赖性定位。

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摘要

The centrosome is the main microtubule-organizing center and constitutes the largest protein complex in a eukaryotic cell. The Dictyostelium centrosome is an established model for acentriolar centrosomes and it consists of a layered core structure surrounded by a so-called corona, which harbors microtubule nucleation complexes. We have identified 34 new centrosomal candidate proteins through mass spectrometrical analysis of the proteome of isolated Dictyostelium centrosomes. Here we present a characterization of 12 centrosomal candidate proteins all featuring coiled coil regions and low expression levels, which are the most common attributes of centrosomal proteins. We used GFP fusion proteins to localize the candidate proteins in whole cells and on microtubule-free, isolated centrosomes. Thus we were able to identify nine new genuine centrosomal proteins including a putative orthologue of Cep192, an interaction partner of polo-like kinase 4 in human centriole biogenesis. In this respect, centrosomal localization of the only polo-like kinase in Dictyostelium, Plk, is also shown in this work. Using confocal deconvolution microscopy, four components, CP39, CP55, CP75, and CP91 could be clearly assigned to the so far almost uncharacterized centrosomal core structure, while CP148 and Cep192 localized to a zone between that of corona marker and core proteins. Finally, CP103 and CP248 were constituents of the corona. In contrast, NE81 was localized at the nuclear envelope and three others, an orthologue of the spindle checkpoint component Mad1, the novel Cenp68, and the centrosomal CP248 were observed at the centromeres, which are clustered and linked to the centrosome throughout the entire cell cycle.
机译：中心体是主要的微管组织中心，并构成真核细胞中最大的蛋白质复合物。 Dictyostelium中心体是无角膜中心体的成熟模型，它由被所谓的电晕所包围的分层核心结构组成，该结构具有微管成核复合物。我们通过质谱分析分离的单壁铁单胞菌中心体的蛋白质组，鉴定出34种新的中心体候选蛋白。在这里，我们介绍了12个中心体候选蛋白的特征，这些蛋白均具有卷曲的螺旋区域和低表达水平，这是中心体蛋白的最常见属性。我们使用GFP融合蛋白将候选蛋白定位在整个细胞中以及无微管的分离的中心体上。因此，我们能够鉴定出九种新的真正的中心体蛋白，包括推定的Cep192直向同源物，Cep192是人类中心体生物发生中polo样激酶4的相互作用伴侣。在这方面，这项工作还显示了盘基网柄菌中唯一的polo样激酶Plk的中心体定位。使用共聚焦反褶积显微镜，可以清楚地将四个组分CP39，CP55，CP75和CP91分配到迄今为止几乎未表征的中心体核心结构，而将CP148和Cep192定位在电晕标记和核心蛋白之间的区域。最后，CP103和CP248是电晕的组成部分。相比之下，NE81位于核被膜上，另外三个位于纺锤体检查点成分Mad1的直向同源物，新型Cenp68和中心体CP248在着丝粒处观察到，它们在整个细胞周期内聚集并与中心体相连。

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