Department of Physics and Astronomy, and Institute of Applied Physics, Seoul National University, Seoul 08826, Republic of Korea.
Department of Chemistry, Korea Advanced Institute of Science and Technology, Daejeon 34141, Republic of Korea.
Nucleic Acids Res. 2023 Apr 11;51(6):2778-2789. doi: 10.1093/nar/gkad051.
Transcriptional pause is essential for all types of termination. In this single-molecule study on bacterial Rho factor-dependent terminators, we confirm that the three Rho-dependent termination routes operate compatibly together in a single terminator, and discover that their termination efficiencies depend on the terminational pauses in unexpected ways. Evidently, the most abundant route is that Rho binds nascent RNA first and catches up with paused RNA polymerase (RNAP) and this catch-up Rho mediates simultaneous releases of transcript RNA and template DNA from RNAP. The fastest route is that the catch-up Rho effects RNA-only release and leads to 1D recycling of RNAP on DNA. The slowest route is that the RNAP-prebound stand-by Rho facilitates only the simultaneous rather than sequential releases. Among the three routes, only the stand-by Rho's termination efficiency positively correlates with pause duration, contrary to a long-standing speculation, invariably in the absence or presence of NusA/NusG factors, competitor RNAs or a crowding agent. Accordingly, the essential terminational pause does not need to be long for the catch-up Rho's terminations, and long pauses benefit only the stand-by Rho's terminations. Furthermore, the Rho-dependent termination of mgtA and ribB riboswitches is controlled mainly by modulation of the stand-by rather than catch-up termination.
转录暂停对于所有类型的终止都是必不可少的。在这项关于细菌 Rho 因子依赖性终止子的单分子研究中，我们证实了三种 Rho 依赖性终止途径可以在单个终止子中共存并协同工作，并且发现它们的终止效率以出人意料的方式依赖于终止暂停。显然，最丰富的途径是 Rho 首先结合新生 RNA，并追上暂停的 RNA 聚合酶 (RNAP)，这种追上的 Rho 介导转录 RNA 和模板 DNA 同时从 RNAP 释放。最快的途径是追上的 Rho 仅导致 RNA 释放，并导致 RNAP 在 DNA 上进行 1D 循环。最慢的途径是 RNAP 预结合的备用 Rho 仅促进同时而不是顺序释放。在这三种途径中，只有备用 Rho 的终止效率与暂停时间呈正相关，这与长期以来的推测相反，无论是否存在 NusA/NusG 因子、竞争 RNA 或拥挤剂。因此，对于追上的 Rho 的终止，必需的终止暂停不需要很长，而长暂停仅有利于备用 Rho 的终止。此外，mgtA 和 ribB 核糖开关的 Rho 依赖性终止主要受到备用终止而不是追上终止的调节。
