RNA degradosomes exist in vivo in Escherichia coli as multicomponent complexes associated with the cytoplasmic membrane via the N-terminal region of ribonuclease E
RNA degradosomes exist in vivo in Escherichia coli as multicomponent complexes associated with the cytoplasmic membrane via the N-terminal region of ribonuclease E
复制标题
DOI：
10.1073/pnas.011535498
复制
发表时间：
2001-01-02
影响因子：
11.1
通讯作者：
Lin-Chao, S
中科院分区：
文献类型：
--
作者：
Liou, GG;Jane, WN;Lin-Chao, S
从大肠杆菌中分离出的 RNase E 包含在多组分“降解体”复合物中，该复合物与参与 RNA 降解的其他蛋白质一起存在。早期的工作表明，RNase E 的 C 末端区域是降解体成分结合的支架，并已鉴定出其与多核苷酸磷酸化酶 (PNPase)、RhlB RNA 解旋酶和烯醇化酶相互作用所必需的特定 RNase E 片段。在这里，我们报告了使用免疫金标记和冷冻断裂方法的电子显微镜研究，表明降解体作为多组分结构存在于大肠杆菌体内，通过 RNase E 的 N 末端区域与细胞质膜结合。虽然 PNPase 和烯醇酶在大肠杆菌中存在，但相对于 RNase E 而言大量过量，因此在细胞中检测到的大部分是与 RNase E 支架无关的分子，免疫金标记和生化分析表明，解旋酶在所有细胞生长阶段，其存在量与 RNase E 大致等摩尔。我们的研究结果确定了大肠杆菌体内基于 RNase E 的降解体的存在和细胞位置，也表明 RNA 加工和降解可能发生在细胞内的特定位点。
RNase E isolated from Escherichia coli is contained in a multicomponent "degradosome" complex with other proteins implicated in RNA decay. Earlier work has shown that the C-terminal region of RNase E is a scaffold for the binding of degradosome components and has identified specific RNase E segments necessary for its interaction with polynucleotide phosphorylase (PNPase), RhlB RNA helicase, and enolase. Here, we report electron microscopy studies that use immunogold labeling and freeze-fracture methods to show that degradosomes exist in vivo in E. coli as multicomponent structures that associate with the cytoplasmic membrane via the N-terminal region of RNase E. Whereas PNPase and enolase are present in E, coli in large excess relative to RNase E and therefore are detected in cells largely as molecules unlinked to the RNase E scaffold, immunogold labeling and biochemical analyses show that helicase is present in approximately equimolar amounts to RNase E at all cell growth stages. Our findings, which establish the existence and cellular location of RNase E-based degradosomes in vivo in E. coli, also suggest that RNA processing and decay may occur at specific sites within cells.