The two steps of Poly(A)-dependent termination, pausing and release, can be uncoupled by truncation of the RNA polymerase II carboxyl-terminal repeat domain
The two steps of Poly(A)-dependent termination, pausing and release, can be uncoupled by truncation of the RNA polymerase II carboxyl-terminal repeat domain
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DOI：
10.1128/mcb.24.10.4092-4103.2004
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发表时间：
2004-05-01
影响因子：
5.3
通讯作者：
Martinson, HG
中科院分区：
文献类型：
--
作者：
Park, NJ;Tsao, DC;Martinson, HG
RNA聚合酶11的羧基末端重复结构域（CTD）被认为有助于协调RNA代谢过程中的事件。哺乳动物CTD由52个不完全重复的七肽组成，随后在C末端有10个额外的残基。CTD是体外切割和聚腺苷酸化所必需的。我们使用CTD截短突变体研究了体内poly（A）依赖性终止。Poly（A）依赖性终止发生在两个步骤中，暂停和释放。我们发现，CTD是释放所必需的，前25个七元组就足够了。不需要最后10个氨基酸，也不需要CTD后半部分的变体七肽。CTD的任何部分都不需要poly（A）依赖性暂停-poly（A）信号可以直接与聚合酶体通信。通过移除CTD，可以观察到暂停，而不会因释放而变得模糊。Poly（A）依赖性暂停似乎通过减慢聚合酶来起作用，例如通过下调正延伸因子。尽管前25个七肽支持未减弱的聚（A）依赖性终止，但它们不能有效地支持参与失败延伸的启动子附近的事件。然而，CTD的后半部分，包括最后10个氨基酸，足以实现这些功能。
The carboxyl-terminal repeat domain (CTD) of RNA polymerase 11 is thought to help coordinate events during RNA metabolism. The mammalian CTD consists of 52 imperfectly repeated heptads followed by 10 additional residues at the C terminus. The CTD is required for cleavage and polyadenylation in vitro. We studied poly(A)-dependent termination in vivo using CTD truncation mutants. Poly(A) -dependent termination occurs in two steps, pause and release. We found that the CTD is required for release, the first 25 heptads being sufficient. Neither the final 10 amino acids nor the variant heptads of the second half of the CTD were required. No part of the CTD was required for poly(A) -dependent pausing-the poly(A) signal could communicate directly with the body of the polymerase. By removing the CTD, pausing could be observed without being obscured by release. Poly (A) -dependent pausing appeared to operate by slowing down the polymerase, such as by down-regulation of a positive elongation factor. Although the first 25 heptads supported undiminished poly(A) -dependent termination, they did not efficiently support events near the promoter involved in abortive elongation. However, the second half of the CTD, including the final 10 amino acids, was sufficient for these functions.