Identification and characterization of proteins involved in the regulation of centrosome duplication

我们研究计划的长期目标是了解中心体生物发生和复制的调控机制。中心体是大多数真核细胞中主要的微管成核中心，在多种生物过程中起着重要作用，包括在细胞分裂过程中有丝分裂纺锤体的建立。虽然中心体在一百多年前就被发现了，但关于它们在细胞周期中的起源和复制的知识仍然缺乏。该细胞器的忠实繁殖是染色体分离的必要条件，中心体复制异常可引起细胞分裂错误，导致细胞突变、生长抑制和细胞死亡。我之前发现了一种新的中心体因子Cep76，它具有独特的功能：它通过特异性地将这一过程限制在每个细胞周期一次来保护中心体复制的保真度。为了进一步描述Cep76调控复制的复杂机制，最近对与Cep76相关的免疫沉淀蛋白进行了蛋白质组学分析，以鉴定新的相互作用蛋白。初步的质谱测序数据揭示了已知的cep76相互作用伙伴，以及中心体复制所必需的蛋白质。此外，还分离到了两个功能未知或部分已知的新蛋白Cep70和Cep78。我们假设Cep70和Cep78与Cep76一样，是中心体复制的重要调节因子。通过两个特定的目标，我们将破译Cep70和Cep78的生物学功能，描绘它们与Cep76的精确关系，并表征它们的蛋白质相互作用网络。这些分析不仅将确定Cep70和Cep78在中心体中的作用，还将确定中心体复制调节所需的关键蛋白质-蛋白质相互作用。预计这些研究结果将进一步阐明中心体复制的分子机制以及该细胞器在细胞增殖和存活中的作用。

The long-term objective of our research program is to understand the regulatory mechanisms underlying the biogenesis and duplication of centrosomes. Centrosomes are the major microtubule-nucleating centers in most eukaryotic cells which are important for a wide variety of biological processes, including the establishment of a mitotic spindle during cell division. Although centrosomes were identified more than a hundred years ago, a paucity of knowledge exists regarding their origins and duplication during the cell cycle. Faithful reproduction of this organelle is essential for chromosome segregation, and abnormalities in centrosome duplication could induce errors in cell division, resulting in cellular catastrophe, growth inhibition, and cell death. I previously identified a novel centrosomal factor, Cep76, with a unique function: it safeguards the fidelity of centrosome duplication by specifically limiting this process to once per cell cycle. To further delineate the intricate mechanisms by which Cep76 regulates duplication, proteomic analysis of immunoprecipitated proteins associated with Cep76 were recently performed to identify novel interacting proteins. Preliminary mass spectrometry sequencing data revealed known Cep76-interacting partners, along with proteins that are essential for centrosome duplication. In addition, two novel proteins of unknown or partially known function, Cep70 and Cep78, were isolated. We hypothesize that Cep70 and Cep78, like Cep76, are important regulators of centrosome duplication. Through two specific aims, we will decipher the biological functions of Cep70 and Cep78, delineate their precise relationship with Cep76, and characterize their protein interaction networks. These analyses will not only define the roles of Cep70 and Cep78 at the centrosome, but will also identify key protein-protein interactions required for the regulation of centrosome duplication. It is anticipated that these study results will further illuminate the molecular mechanisms underlying centrosome duplication and the role of this organelle in cell proliferation and survival.