Visual Analyses of Long Single-stranded RNA Genome

Research Project

Project/Area Number	18K06082
Research Category	Grant-in-Aid for Scientific Research (C)
Allocation Type	Multi-year Fund
Section	一般
Review Section	Basic Section 43020:Structural biochemistry-related
Research Institution	Kyoto University
Principal Investigator	Takeyasu Kunio 京都大学, 生命科学研究科, 研究員 (40135695)
Project Period (FY)	2018-04-01 – 2021-03-31
Project Status	Completed (Fiscal Year 2020)
Budget Amount *help	¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000) Fiscal Year 2020: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000) Fiscal Year 2019: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000) Fiscal Year 2018: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Keywords	長い一本鎖RNAの高次構造 / RNAウィルスゲノム / 原子間力顕微鏡 / RNAの高次構造 / HCVゲノムRNAの高次構造 / 原子間力顕微鏡法（AFM) / 長い一本鎖RNAの高次構造解析 / 原子間力現鼻鏡法（AFM) / RNAゲノムフォールディング / RNA genome structure / Atomic force microscopy / Singe-stranded RNA
Outline of Final Research Achievements	Structural analyses of long single-stranded RNA (~10 kb) have been very difficult. For example, it took 20 some years to determine the secondary structure of 28S rRNA using NMR and X-ray crystallography. It is impossible to acquire such secondary structures of much longer single-stranded RNAs such as RNA virus entire genome. We have been applying Atomic Force Microscopy (AFM) to the structural analyses of HCV genome RNA. The outcome of our investigation showed that the HCV genome can be spread out nicely on mica surface exhibiting main chain with many small domains branching out of the main chain. These domains consist of Watson-Crick-type double stranded structures. The number of nucleotides in each domain can be estimated from the volume the domain. Thus, from these data, a secondary structural model of entire HCV genome can be constructed.
Academic Significance and Societal Importance of the Research Achievements	［長い一本鎖RNAのとる高次構造は如何に］この問いに答えることは、以下のような究極的問題の解決へのカギとなろう。（１）RNAウィルスのゲノム構造構築・パッケージング機構（２）翻訳におけるmRNAの高次構造と翻訳調節、翻訳効率との関係（３）RNAの安定性と高次構造との関係、それに及ぼす結合タンパク質の影響。学術的には、“まるごとから見た”長い一本鎖RNAウィルスゲノムの高次構造（特に二次構造とそれへのRNA結合タンパク質の影響）への挑戦であり、将来の「ウィルスゲノムのパッケージング機構の解明」にもつながる。方法論的には、“Simple”であり、分子生物学と構造生物学との融合である。