Department of Biophysics, Johns Hopkins University, Baltimore, MD, 21218, USA.
Physics Frontier Center (Center for Physics of Living Cells), University of Illinois, Urbana, IL, 61801, USA.
Nat Commun. 2024 May 10;15(1):3963. doi: 10.1038/s41467-024-48247-8.
Translation initiation in bacteria is frequently regulated by various structures in the 5' untranslated region (5'UTR). Previously, we demonstrated that G-quadruplex (G4) formation in non-template DNA enhances transcription. In this study, we aim to explore how G4 formation in mRNA (RG4) at 5'UTR impacts translation using a T7-based in vitro translation system and in E. coli. We show that RG4 strongly promotes translation efficiency in a size-dependent manner. Additionally, inserting a hairpin upstream of the RG4 further enhances translation efficiency, reaching up to a 12-fold increase. We find that the RG4-dependent effect is not due to increased ribosome affinity, ribosome binding site accessibility, or mRNA stability. We propose a physical barrier model in which bulky structures in 5'UTR biases ribosome movement toward the downstream start codon, thereby increasing the translation output. This study provides biophysical insights into the regulatory role of 5'UTR structures in in vitro and bacterial translation, highlighting their potential applications in tuning gene expression.
在细菌中，翻译起始通常受到 5'非翻译区（5'UTR）中各种结构的调节。之前，我们证明了非模板 DNA 中的 G-四链体（G4）形成可增强转录。在这项研究中，我们旨在使用 T7 体外翻译系统和大肠杆菌来探索 5'UTR 中 mRNA（RG4）中的 G4 形成如何影响翻译。我们发现 RG4 以大小依赖的方式强烈促进翻译效率。此外，在 RG4 上游插入发夹结构进一步增强了翻译效率，最高可达 12 倍的增加。我们发现，RG4 依赖性效应不是由于增加了核糖体亲和力、核糖体结合位点的可及性或 mRNA 的稳定性。我们提出了一个物理障碍模型，其中 5'UTR 中的大体积结构会使核糖体向下游起始密码子移动，从而增加翻译产量。这项研究为 5'UTR 结构在体外和细菌翻译中的调节作用提供了生物物理见解，突出了它们在调节基因表达方面的潜在应用。
