Institute of Cell Biology, University of Bern, Bern, Switzerland.
Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland.
PLoS Negl Trop Dis. 2020 Sep 16;14(9):e0008568. doi: 10.1371/journal.pntd.0008568. eCollection 2020 Sep.
Trypanosoma brucei is a single celled eukaryotic parasite and the causative agent of human African trypanosomiasis and nagana in cattle. Aside from its medical relevance, T. brucei has also been key to the discovery of several general biological principles including GPI-anchoring, RNA-editing and trans-splicing. The parasite contains a single mitochondrion with a singular genome. Recent studies have identified several molecular components of the mitochondrial genome segregation machinery (tripartite attachment complex, TAC), which connects the basal body of the flagellum to the mitochondrial DNA of T. brucei. The TAC component in closest proximity to the mitochondrial DNA is TAC102. Here we apply and compare three different approaches (proximity labelling, immunoprecipitation and yeast two-hybrid) to identify novel interactors of TAC102 and subsequently verify their localisation. Furthermore, we establish the direct interaction of TAC102 and p166 in the unilateral filaments of the TAC.
布氏锥虫是一种单细胞真核寄生虫，也是引起人类非洲锥虫病和牛纳格病的病原体。除了具有医学意义外，布氏锥虫还为发现几个一般生物学原理提供了关键线索，包括 GPI 锚定、RNA 编辑和转剪接。寄生虫只有一个带有单一基因组的线粒体。最近的研究已经确定了线粒体基因组分离机制（三分体附着复合物，TAC）的几个分子组成部分，该机制将鞭毛的基底连接到布氏锥虫的线粒体 DNA。与线粒体 DNA 最接近的 TAC 组件是 TAC102。在这里，我们应用并比较了三种不同的方法（邻近标记、免疫沉淀和酵母双杂交）来鉴定 TAC102 的新相互作用蛋白，并随后验证它们的定位。此外，我们还建立了 TAC102 和 p166 在 TAC 的单侧丝状体中的直接相互作用。
