Dynamic Accommodation of mRNA onto Ribosomes Facilitates Translation Initiation

• Main Authors: Yi-Lan Chen, 陳臆嵐
• Other Authors: Jin-Der Wen
• Format: Others
• Language: en_US
• Published: 2019
• Online Access: http://ndltd.ncl.edu.tw/handle/ejpzun

博士 === 國立臺灣大學 === 基因體與系統生物學學位學程 === 107 === Regulation of translation initiation is important for protein synthesis and protein homeostasis in prokaryotic cells. During initiation, the ribosome encounters mRNAs with different sequences and structures. Therefore, understanding how different mRNAs correctly accommodate the ribosome and maintain the fidelity of translation at the beginning of initiation is essential. In this study, by directly observing the 30S ribosomal subunit and mRNA interaction by single-molecule FRET, we found the mRNA wraps the small subunit back and forth dynamically. This movement becomes more frequent if the downstream sequence forms structures, such as duplexes and hairpins. After measuring the unfolding force of downstream structures, we found the mRNA movement during initiation helps the ribosome destabilize downstream structures. We suggest that this dynamic movement helps the ribosome to search and find correct initiation site. Interaction between mRNA and the ribosome through strong Shine-Dalgarno sequence prevents the ribosome from dropping from the mRNA during the dynamic movement. Addition of initiator tRNA forms the 30S initiation complex (30S IC). Here, initiator tRNA suppresses the dynamic movement and opens downstream structures up to 3 bp. This step is a checkpoint for the correct initiation. Initiation factors, especially IF3, also stabilizes the mRNA before and after the initiator tRNA anchoring. However, IF3 also locally destabilize the 30S IC. However, wrong stable 30S IC may form, especially when the downstream structure is a duplex structure. The duplex structures can be formed through small RNA (sRNA). In this case, IF3 can rescue the complex by removing the initiator tRNA and dissociating the mRNA-ribosome complex. Here, we reveal that the flexibility of mRNA movement, the selectivity of tRNA decoding, and the assurance of initiation factors help the ribosome maintain the robustness and fidelity of translation at the early initiation stage.