On the stability of stalled RNA polymerase and its removal by RapA.
On the stability of stalled RNA polymerase and its removal by RapA.
复制标题
DOI：
10.1093/nar/gkac558
复制
发表时间：
2022-07-22
影响因子：
14.9
通讯作者：
Strick, Terence R.
中科院分区：
文献类型：
--
作者：
Portman, James R.;Qayyum, M. Zuhaib;Murakami, Katsuhiko S.;Strick, Terence R.
由DNA、RNA聚合酶（RNAP）和RNA形成的转录延伸复合物的停滞由于DNA结合的聚合酶的极高稳定性而对并行过程造成严重障碍。已知RapA以ATP依赖性方式从DNA中去除RNAP，在50多年前被鉴定为RNAP的丰富结合伴侣;然而，其作用机制仍然未知。在这里，我们使用单分子磁捕获测定来表征RapA活性，并开始指定其作用机制。我们首先表明，停滞RNAP驻留在DNA上的时间为106秒的顺序，并增加DNA上的正扭矩减少了这个寿命。使用停滞的RNAP作为底物，我们表明，RapA蛋白刺激停滞的RNAP从正超螺旋DNA，但不是负超螺旋DNA的解离。我们观察到，RapA依赖的RNAP解离是扭矩敏感的，抑制GreB和依赖于RNA长度。我们建议，停滞RNAP是从DNA中驱逐RapA通过回溯在一个超螺旋和扭矩依赖的方式，这表明RapA的活性在体内转录RNAP负责解决聚合酶分子马达之间的冲突。
Stalling of the transcription elongation complex formed by DNA, RNA polymerase (RNAP) and RNA presents a serious obstacle to concurrent processes due to the extremely high stability of the DNA-bound polymerase. RapA, known to remove RNAP from DNA in an ATP-dependent fashion, was identified over 50 years ago as an abundant binding partner of RNAP; however, its mechanism of action remains unknown. Here, we use single-molecule magnetic trapping assays to characterize RapA activity and begin to specify its mechanism of action. We first show that stalled RNAP resides on DNA for times on the order of 106 seconds and that increasing positive torque on the DNA reduces this lifetime. Using stalled RNAP as a substrate we show that the RapA protein stimulates dissociation of stalled RNAP from positively supercoiled DNA but not negatively supercoiled DNA. We observe that RapA-dependent RNAP dissociation is torque-sensitive, is inhibited by GreB and depends on RNA length. We propose that stalled RNAP is dislodged from DNA by RapA via backtracking in a supercoiling- and torque-dependent manner, suggesting that RapA’s activity on transcribing RNAP in vivo is responsible for resolving conflicts between converging polymerase molecular motors.
登录
查看更多内容
RNA聚合酶扭矩产生能力的转录因子调节
DOI：
10.1073/pnas.1807031116
发表时间：
2019-02-12
影响因子：
11.1
作者：
Ma, Jie;Tan, Chuang;Wang, Michelle D.
通讯作者：
Wang, Michelle D.
DOI：
10.1073/pnas.2019630118
发表时间：
2021-04-13
影响因子：
11.1
作者：
Portman JR;Brouwer GM;Bollins J;Savery NJ;Strick TR
通讯作者：
Strick TR
影响因子：
64.8
作者：
BURGESS, RR;TRAVERS, AA;BAUTZ, EKF
通讯作者：
BAUTZ, EKF
影响因子：
11.4
作者：
Toulmé, F;Mosrin-Hauman, C;Rahmouni, AR
通讯作者：
Rahmouni, AR
影响因子：
14.9
作者：
McKinley, Brian A;Sukhodolets, Maxim V
通讯作者：
Sukhodolets, Maxim V