CENTRIOLE DUPLICATION IN C ELEGANS EMBRYOS

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。中心粒是纤毛和鞭毛形成的重要细胞器，参与细胞质分裂、细胞周期控制和中心体组织。细胞的单中心体作为纺锤杆遗传，每个细胞周期只复制一次，这一过程是通过控制中心粒复制来调节的。中心粒复制受到严格控制，以确保双极纺锤体的形成和倍性的维持。多年来人们已经知道，子中心粒与老中心粒或“母中心粒”成直角组装。这一发现的意义和子中心粒聚集的途径仍然是个谜。我们正在使用子项目0067中描述的相关光和EM断层扫描方法来研究分阶段秀丽隐杆线虫胚胎中的中心粒复制。我们的初步结果表明，这种生物体中的子中心粒组装始于中心管的形成，就像原核出现一样。中心管在原核迁移过程中伸长，单线态微管在原核旋转过程中聚集。通过em断层扫描结合RNA干扰和恢复显微镜，我们发现了证据表明，试管的形成和伸长需要SAS-5/SAS-6蛋白和由SPD-2和ZYG-1介导的上游信号。我们进一步表明，尽管中心管的组装仍然发生，但单线态微管在没有SAS-4的情况下无法组装，这表明在组装的这一步特别需要SAS-4。我们在线虫中描述的中心粒组装途径可能为进一步研究其他携带中心粒的生物的中心粒组装提供一个框架。这部作品正在写完准备出版。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Centrioles are organelles that play essential roles in the formation of cilia and flagella, and they participate in cytokinesis, cell cycle control and centrosome organization. The cell¿s single centrosome, which is inherited as a spindle pole, duplicates only once per cell cycle, a process that is regulated through the control of centriole duplication. Centriole duplication is strictly controlled to ensure the formation of a bipolar spindle and the maintenance of ploidy. It has been know for many years that daughter centrioles assemble at right angles to the older, or ¿mother¿ centriole. The significance of this and the pathway for daughter centriole assemble remain mysterious. We are using the correlative light and EM tomography approach described in Subproject 0067 to study centriole duplication in staged C. elegans embryos. Our initial results showed that daughter centriole assembly in this organism begins with the formation of a central tube, just as the pronuclei appear. The central tube elongates during pronuclear migration, and the assembly of singlet microtubules on the tube occurs during pronuclear rotation. Using EM-tomography in combination with RNA interference and restoration microscopy we have found evidence suggesting that tube formation and elongation requires the SAS-5/SAS-6 proteins and an upstream signal mediated by SPD-2 and ZYG-1. We further show that although the assembly of the central tube still occurs, singlet microtubules fail to assemble in the absence of SAS-4, suggesting that it is required specifically at that step of assembly. The centriole assembly pathway we describe here in C. elegans may provide a framework for further studies on centriole assembly in other centriole bearing organisms. This work is being written up for publication.