Productive mRNA stem loop-mediated transcriptional slippage: Crucial features in common with intrinsic terminators
Productive mRNA stem loop-mediated transcriptional slippage: Crucial features in common with intrinsic terminators
复制标题
DOI：
10.1073/pnas.1418384112
复制
发表时间：
2015-04-21
影响因子：
11.1
通讯作者：
Atkins, John F.
中科院分区：
文献类型：
--
作者：
Penno, Christophe;Sharma, Virag;Atkins, John F.
大肠杆菌和酵母 DNA 依赖性 RNA 聚合酶可介导有效的新生转录物茎环形成依赖性 RNA-DNA 杂合重排。在异聚序列 T5C5 上发现了重排，并产生了相应 U5C4 内缺少 C 残基的转录本。研究的序列源自 Roseiflexus 插入序列 (IS) 元件，其中产生的转录滑移是转座酶合成所必需的。 RNA 结构的稳定性、茎环与滑动位点的接近度、滑动位点基序的长度和组成以及其 3' 相邻核苷酸 (nt) 的身份对于缺少单个 C 的转录物至关重要。在许多方面，这种滑动的 RNA 结构要求类似于发夹依赖性转录终止的要求。在纯化的体外系统中，滑动效率范围为 5% 至 75%，具体取决于滑动序列和 3'nt 上下文指定的核苷酸的浓度比。之前关于茎环介导的滑动的唯一提议（在埃博拉病毒表达中）是基于不正确的数据解释。我们提出了一种机械滑移模型，涉及 RNAP 易位状态作为滑移方向性和效率的主要动力。它与之前描述的模型不同，包括为副粘病毒提出的模型，其中随机运动效率主要取决于新的重新排列的杂交体的稳定性。在扩大转录滑移用于基因表达的利用范围时，刺激结构在翻译水平上提供了与程序化核糖体移码的相似之处。
Escherichia coli and yeast DNA-dependent RNA polymerases are shown to mediate efficient nascent transcript stem loop formation-dependent RNA-DNA hybrid realignment. The realignment was discovered on the heteropolymeric sequence T5C5 and yields transcripts lacking a C residue within a corresponding U5C4. The sequence studied is derived from a Roseiflexus insertion sequence (IS) element where the resulting transcriptional slippage is required for transposase synthesis. The stability of the RNA structure, the proximity of the stem loop to the slippage site, the length and composition of the slippage site motif, and the identity of its 3' adjacent nucleotides (nt) are crucial for transcripts lacking a single C. In many respects, the RNA structure requirements for this slippage resemble those for hairpin-dependent transcription termination. In a purified in vitro system, the slippage efficiency ranges from 5% to 75% depending on the concentration ratios of the nucleotides specified by the slippage sequence and the 3' nt context. The only previous proposal of stem loop mediated slippage, which was in Ebola virus expression, was based on incorrect data interpretation. We propose a mechanical slippage model involving the RNAP translocation state as the main motor in slippage directionality and efficiency. It is distinct from previously described models, including the one proposed for paramyxovirus, where following random movement efficiency is mainly dependent on the stability of the new realigned hybrid. In broadening the scope for utilization of transcription slippage for gene expression, the stimulatory structure provides parallels with programmed ribosomal frameshifting at the translation level.