Physiological function of RNase H resolving R-loop structures formed on chromsome

2022 Fiscal Year Final Research Report

• Project/Area Number: 20K15721
• Research Category: Grant-in-Aid for Early-Career Scientists
• Allocation Type: Multi-year Fund
• Review Section: Basic Section 43010:Molecular biology-related
• Research Institution: Aichi Cancer Center Research Institute
• Principal Investigator: Ryo Uehara 愛知県がんセンター(研究所), 腫瘍制御学分野, 研究員 (70842590)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Keywords: RNase H2 / R-loop / 自己免疫疾患 / ゲノム不安定性

Outline of Final Research Achievements

R-loop is a three-stranded structure composed of RNA/DNA heteroduplex and isolated single stranded DNA. RNase H2 is a major enzyme that hydrolases RNA/DNA hybrids in R-loops and prevents DNA damage events caused by persistent R-loops. To investigate the effects on R-loop accumulation, we generated HEK293 cells carrying the mutation that makes RNase H2 inactive on R-loop. Using these cells defective in R-loop resolution, we revealed that R-loop accumulation in HEK293 did not seriously disturb the genome stability. In addition, we developed a synthetic binding protein (SBP), which binds to archaeal RNase H2 and inhibits its RNA/DNA hydrolysis, and determined the crystal structure of RNase H2 and SBP complex. This structure revealed the two loops in active site seemed to be important for hydrolysis activity of RNA/DNA hybrids among archaeal and eukaryotic enzymes.

Free Research Field

分子生物学

Academic Significance and Societal Importance of the Research Achievements

RNase H2は原核生物からヒトに至るまでほぼ全ての生物に存在するが、R-loopを分解する活性はアーキアと真核生物のみが獲得しており、詳細な分子機構や生理的意義が明らかではない。本研究では、アーキア由来の酵素と人工阻害タンパク質の複合体の立体構造からそのメカニズムに迫った。また、RNase H2のサブユニットへの変異は神経変性疾患エカルディ・グティエール症候群（AGS）の原因であるため、R-loopの蓄積に伴う生体応答はAGS発症メカニズムに深く関与することが予想される。RNase H2のR-loop分解機構の解明は、学術・医療の両面において意義深い。