Department of Chemistry, University of North Carolina, Chapel Hill, NC 27599-3290, USA.
Department of Medicine, University of North Carolina, Chapel Hill, NC 27599, USA; UNC HIV Cure Center, University of North Carolina, Chapel Hill, NC 27599, USA.
Mol Cell. 2022 May 5;82(9):1708-1723.e10. doi: 10.1016/j.molcel.2022.02.009. Epub 2022 Mar 22.
7SK is a conserved noncoding RNA that regulates transcription by sequestering the transcription factor P-TEFb. 7SK function entails complex changes in RNA structure, but characterizing RNA dynamics in cells remains an unsolved challenge. We developed a single-molecule chemical probing strategy, DANCE-MaP (deconvolution and annotation of ribonucleic conformational ensembles), that defines per-nucleotide reactivity, direct base pairing interactions, tertiary interactions, and thermodynamic populations for each state in RNA structural ensembles from a single experiment. DANCE-MaP reveals that 7SK RNA encodes a large-scale structural switch that couples dissolution of the P-TEFb binding site to structural remodeling at distal release factor binding sites. The 7SK structural equilibrium shifts in response to cell growth and stress and can be targeted to modulate expression of P-TEFbresponsive genes. Our study reveals that RNA structural dynamics underlie 7SK function as an integrator of diverse cellular signals to control transcription and establishes the power of DANCE-MaP to define RNA dynamics in cells.
7SK 是一种保守的非编码 RNA，通过隔离转录因子 P-TEFb 来调节转录。7SK 的功能需要 RNA 结构的复杂变化，但在细胞中描述 RNA 动力学仍然是一个未解决的挑战。我们开发了一种单分子化学探测策略，DANCE-MaP（核糖核酸构象集合的去卷积和注释），该策略从单个实验中为 RNA 结构集合中的每个状态定义了核苷酸反应性、直接碱基配对相互作用、三级相互作用和热力学种群。DANCE-MaP 表明，7SK RNA 编码了一种大规模的结构开关，该开关将 P-TEFb 结合位点的溶解与远端释放因子结合位点的结构重塑联系起来。7SK 的结构平衡会响应细胞生长和应激而发生变化，并可以作为靶向调节 P-TEFb 反应基因表达的靶点。我们的研究揭示了 7SK 作为整合多种细胞信号以控制转录的功能的基础是 RNA 结构动力学，并确立了 DANCE-MaP 在细胞中定义 RNA 动力学的能力。
