Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, CA 90095, USA.
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA 90095, USA; California NanoSystems Institute, University of California, Los Angeles, Los Angeles, CA 90095, USA.
Mol Cell. 2022 May 5;82(9):1724-1736.e7. doi: 10.1016/j.molcel.2022.03.001. Epub 2022 Mar 22.
7SK non-coding RNA (7SK) negatively regulates RNA polymerase II (RNA Pol II) elongation by inhibiting positive transcription elongation factor b (P-TEFb), and its ribonucleoprotein complex (RNP) is hijacked by HIV-1 for viral transcription and replication. Methylphosphate capping enzyme (MePCE) and La-related protein 7 (Larp7) constitutively associate with 7SK to form a core RNP, while P-TEFb and other proteins dynamically assemble to form different complexes. Here, we present the cryo-EM structures of 7SK core RNP formed with two 7SK conformations, circular and linear, and uncover a common RNA-dependent MePCE-Larp7 complex. Together with NMR, biochemical, and cellular data, these structures reveal the mechanism of MePCE catalytic inactivation in the core RNP, unexpected interactions between Larp7 and RNA that facilitate a role as an RNP chaperone, and that MePCE-7SK-Larp7 core RNP serves as a scaffold for switching between different 7SK conformations essential for RNP assembly and regulation of P-TEFb sequestration and release.
7SK 非编码 RNA（7SK）通过抑制正转录延伸因子 b（P-TEFb）负调控 RNA 聚合酶 II（RNA Pol II）的延伸，其核糖核蛋白复合物（RNP）被 HIV-1 劫持用于病毒转录和复制。甲基磷酸化酶（MePCE）和 La 相关蛋白 7（Larp7）与 7SK 组成性结合形成核心 RNP，而 P-TEFb 和其他蛋白质则动态组装形成不同的复合物。在这里，我们展示了由两种 7SK 构象（环状和线性）形成的 7SK 核心 RNP 的冷冻电镜结构，并揭示了 RNA 依赖性 MePCE-Larp7 复合物的共同结构。结合 NMR、生化和细胞数据，这些结构揭示了核心 RNP 中 MePCE 催化失活的机制、Larp7 与 RNA 之间意想不到的相互作用，这些相互作用有助于其作为 RNP 伴侣的作用，以及 MePCE-7SK-Larp7 核心 RNP 作为切换不同 7SK 构象的支架的作用，这些构象对于 RNP 组装和 P-TEFb 隔离和释放的调控至关重要。
