Howard Hughes Medical Institute, University of Colorado, Boulder, CO.
Department of Biochemistry, University of Colorado, Boulder, CO.
J Cell Biol. 2018 Dec 3;217(12):4124-4140. doi: 10.1083/jcb.201806183. Epub 2018 Oct 15.
Stress granules (SGs) are transient membraneless organelles of nontranslating mRNA-protein complexes (mRNPs) that form during stress. In this study, we used multiple single-molecule FISH probes for particular mRNAs to examine their SG recruitment and spatial organization. Ribosome runoff is required for SG entry, as long open reading frame (ORF) mRNAs are delayed in SG accumulation, indicating that the SG transcriptome changes over time. Moreover, mRNAs are 20× compacted from an expected linear length when translating and compact 2-fold further in a stepwise manner beginning at the 5' end during ribosome runoff. Surprisingly, the 5' and 3' ends of the examined mRNAs were separated when translating, but in nontranslating conditions the ends of long ORF mRNAs become close, suggesting that the closed-loop model of mRNPs preferentially forms on nontranslating mRNAs. Compaction of ribosome-free mRNAs is ATP independent, consistent with compaction occurring through RNA structure formation. These results suggest that translation inhibition triggers an mRNP reorganization that brings ends closer, which has implications for the regulation of mRNA stability and translation by 3' UTR elements and the poly(A) tail.
应激颗粒（SGs）是在应激过程中形成的非翻译 mRNA-蛋白复合物（mRNPs）的瞬态无膜细胞器。在这项研究中，我们使用了多个特定 mRNA 的单分子 FISH 探针来检测它们在 SG 中的募集和空间组织。核糖体流导致 SG 的进入，因为长开放阅读框（ORF）mRNA 在 SG 积累中被延迟，这表明 SG 的转录组随时间发生变化。此外，当翻译时，mRNA 从预期的线性长度压缩 20 倍，并且在核糖体流开始时从 5' 端以逐步方式进一步压缩 2 倍。令人惊讶的是，在翻译时，被检查的 mRNA 的 5' 和 3' 末端被分离，但在非翻译条件下，长 ORF mRNA 的末端变得接近，这表明 mRNP 优先在非翻译的 mRNA 上形成闭环模型。无核糖体的 mRNA 压缩与 ATP 无关，这与通过 RNA 结构形成的压缩一致。这些结果表明，翻译抑制触发了 mRNP 的重组，使末端更接近，这对通过 3'UTR 元件和 poly(A)尾对 mRNA 稳定性和翻译的调控具有重要意义。
