Characterisation of genes that regulate centriole stability and ciliogenesis

在大多数有丝分裂中发生的染色体分离主要是由微管产生的力完成的。在大多数情况下，这些微管来自中心体微管组织中心。中心体是一种复杂的结构，包括一对中心粒，被称为中心粒周围物质的蛋白质包围。在每次有丝分裂细胞分裂时，中心粒对必须分裂、复制并分离到纺锤体的相反两极，这被称为中心体周期。然而，当细胞停止分裂以执行其终末分化程序时，中心粒也必须认识到这种情况并相应地停止中心体周期。哺乳动物上皮细胞在最终分裂后，子细胞中存在的未复制的中心粒发生特殊变化并迁移到细胞膜，在那里它们将产生初级纤毛。这些细胞器是细胞膜的延伸，可以促进细胞运动，机械转导，同时也在细胞信号传导中发挥核心作用。在秀丽隐杆线虫中，纤毛局限于感觉神经元；分化的上皮细胞不具有初级纤毛，也没有中心粒/中心体。我们已经证明，在特定的发育背景下，中心粒从细胞周期中分离，而在肠道、外阴和皮下的上皮细胞中，中心粒在最终的细胞分裂后被主动消除。因此，秀丽隐杆线虫的上皮细胞可能不具有初级纤毛，因为它们的中心粒在终末分化时被主动消除。***为了研究最终分裂后中心粒消除的过程，我们通过遗传分析来识别在分化时不消除中心粒的突变体。使用中心粒特异性标记，我们试图分离干扰分化上皮（即肠）中中心粒及时消除的突变体。从这个试点筛选中，我们确定了一个突变，使中心粒在秀丽隐杆线虫的肠道上皮细胞中持续存在。然而，这些中心粒不会成为纤毛，这表明有丝分裂后中心粒的存在并不足以形成纤毛。我们将使用经典的遗传分析和新的转基因方法来鉴定可能阻止这种转化或将有丝分裂中心粒转化为纤毛所需的基因。

The chromosome segregation that occurs during most mitotic divisions is accomplished largely by forces generated by microtubules. In most cases these microtubules emanate from the centrosome-based microtubule organising centre. The centrosome is a complex structure that comprises a centriole pair surrounded by a proteinaceous nuage referred to as the pericentriolar material. At each mitotic cell division the centriole pair must split, duplicate and separate to opposing poles of the spindle in what is referred to as the centrosome cycle. However when cells cease their divisions to execute their terminal differentiation program the centrioles too must recognize this condition and accordingly arrest the centrosome cycle. Following the final division in mammalian epithelial cells the unduplicated centrioles present in the daughter cells undergo specific changes and migrate to the cell membrane where they will generate the primary cilium. These organelles are extensions of the cell membrane and can contribute to cell motility, mechanotransduction, while also playing a central role in cell signalling. ***In C. elegans, cilia are restricted to the sensory neurons; differentiated epithelial cells do not possess primary cilia nor do they show evidence of a centriole/centrosome. We have shown that in specific developmental contexts the centrioles uncouple from the cell cycle, while in the epithelial cells of the gut, the vulva, and the hypodermis, the centrioles are actively eliminated following the final cell division. The epithelial cells of C. elegans therefore do not possess primary cilia potentially because their centrioles are actively eliminated upon terminal differentiation. ***To investigate the process involved in centriole elimination following the final division we genetic analysis to identify mutants that do not eliminate their centrioles upon differentiation. Using a centriole-specific marker we sought to isolate mutants that disrupted the timely elimination of the centrioles in differentiated epithelia, namely in the intestine. From this pilot screen we identified one mutant that allows the centrioles to persist in the gut epithelial cells in C. elegans. These centrioles however do not become cilia suggesting that the presence of a post-mitotic centriole is not sufficient for the formation of a cilium. We will use by classic genetic analysis and novel transgenic approaches to identify genes that may block this conversion or are required to convert a mitotic centriole into a cilium.