Bateson Centre and the Department of Biomedical Science, University of Sheffield, Western Bank, Sheffield, S10 2TN, UK.
Nencki Institute of Experimental Biology Polish Academy of Sciences - Laboratory of Cytoskeleton and Cilia Biology, 3 Pasteur Street, 02-093, Warsaw, Poland.
Sci Rep. 2021 Apr 12;11(1):7899. doi: 10.1038/s41598-021-86909-5.
The base of the cilium comprising the transition zone (TZ) and transition fibers (TF) acts as a selecting gate to regulate the intraflagellar transport (IFT)-dependent trafficking of proteins to and from cilia. Before entering the ciliary compartment, IFT complexes and transported cargoes accumulate at or near the base of the cilium. The spatial organization of IFT proteins at the cilia base is key for understanding cilia formation and function. Using stochastic optical reconstruction microscopy (STORM) and computational averaging, we show that seven TZ, nine IFT, three Bardet-Biedl syndrome (BBS), and one centrosomal protein, form 9-clustered rings at the cilium base of a ciliate Tetrahymena thermophila. In the axial dimension, analyzed TZ proteins localize to a narrow region of about 30 nm while IFT proteins dock approximately 80 nm proximal to TZ. Moreover, the IFT-A subcomplex is positioned peripheral to the IFT-B subcomplex and the investigated BBS proteins localize near the ciliary membrane. The positioning of the HA-tagged N- and C-termini of the selected proteins enabled the prediction of the spatial orientation of protein particles and likely cargo interaction sites. Based on the obtained data, we built a comprehensive 3D-model showing the arrangement of the investigated ciliary proteins.
纤毛的基体包括过渡区（TZ）和过渡纤维（TF），作为一个选择门控，调节依赖于鞭毛内运输（IFT）的蛋白向纤毛和从纤毛的运输。在进入纤毛室之前，IFT 复合物和被运输的货物在纤毛的基部积累或靠近纤毛的基部。IFT 蛋白在纤毛基体的空间组织对于理解纤毛的形成和功能是关键的。我们使用随机光学重建显微镜（STORM）和计算平均法，显示了在纤毛虫四膜虫的纤毛基体处，七个 TZ、九个 IFT、三个 Bardet-Biedl 综合征（BBS）和一个中心体蛋白形成 9 个簇状环。在轴向维度上，分析的 TZ 蛋白定位于大约 30nm 的狭窄区域，而 IFT 蛋白在 TZ 附近约 80nm 处对接。此外，IFT-A 亚基复合物定位于 IFT-B 亚基复合物的外围，并且所研究的 BBS 蛋白位于纤毛膜附近。所选蛋白的 HA 标记的 N-和 C-末端的定位使得能够预测蛋白颗粒和可能的货物相互作用位点的空间取向。基于所获得的数据，我们构建了一个综合的 3D 模型，显示了所研究的纤毛蛋白的排列。
