Department of Biochemistry, McGill University, Montreal, QC H3G 0B1, Canada; Centre de recherche en biologie structurale (CRBS), McGill University, Montreal, QC H3G 0B1, Canada.
Department of Microbiology, Cornell University, Ithaca, NY 14853, USA.
Mol Cell. 2024 Jun 20;84(12):2368-2381.e6. doi: 10.1016/j.molcel.2024.05.012. Epub 2024 Jun 3.
The Tn7 family of transposons is notable for its highly regulated integration mechanisms, including programmable RNA-guided transposition. The targeting pathways rely on dedicated target selection proteins from the TniQ family and the AAA+ adaptor TnsC to recruit and activate the transposase at specific target sites. Here, we report the cryoelectron microscopy (cryo-EM) structures of TnsC bound to the TniQ domain of TnsD from prototypical Tn7 and unveil key regulatory steps stemming from unique behaviors of ATP- versus ADP-bound TnsC. We show that TnsD recruits ADP-bound dimers of TnsC and acts as an exchange factor to release one protomer with exchange to ATP. This loading process explains how TnsC assembles a heptameric ring unidirectionally from the target site. This unique loading process results in functionally distinct TnsC protomers within the ring, providing a checkpoint for target immunity and explaining how insertions at programmed sites precisely occur in a specific orientation across Tn7 elements.
Tn7 家族转座子以其高度调控的整合机制而闻名，包括可编程 RNA 指导的转座。靶向途径依赖于来自 TniQ 家族的专用靶标选择蛋白和 AAA+接头 TnsC，以在特定靶标位点募集和激活转座酶。在这里，我们报告了与源自典型 Tn7 的 TnsD 的 TniQ 结构域结合的 TnsC 的冷冻电镜 (cryo-EM) 结构，并揭示了源自 ATP 与 ADP 结合的 TnsC 的独特行为的关键调节步骤。我们表明，TnsD 招募 ADP 结合的 TnsC 二聚体，并作为交换因子释放一个与 ATP 交换的单体。这个加载过程解释了 TnsC 如何从靶位点单向组装一个七聚体环。这种独特的加载过程导致环内的 TnsC 单体具有不同的功能，为靶标免疫提供了一个检查点，并解释了为什么在编程位点的插入能够以特定的方向精确地发生在 Tn7 元件中。
