2021 Fiscal Year Final Research Report

A multidisciplinary approach to establish the structural basis of dosage compensation

Research Project

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Project/Area Number	19H03169
Research Category	Grant-in-Aid for Scientific Research (B)
Allocation Type	Single-year Grants
Section	一般
Review Section	Basic Section 43020:Structural biochemistry-related
Research Institution	Tokyo University of Pharmacy and Life Science (2021) Tokyo Metropolitan University (2019-2020)
Principal Investigator	Mishima Masaki 東京薬科大学, 薬学部, 教授 (70346310)
Co-Investigator(Kenkyū-buntansha)	坂本泰一千葉工業大学, 先進工学部, 教授 (40383369) 田岡万悟東京都立大学, 理学研究科, 准教授 (60271160) 藤原俊伸近畿大学, 薬学部, 教授 (80362804)
Project Period (FY)	2019-04-01 – 2022-03-31
Keywords	遺伝子量補償 / SHARP / NMR / X線結晶構造解析
Outline of Final Research Achievements	Xist, an lncRNA, plays a pivotal role in global transcriptional repression at the chromosomal level in dosage compensation. To understand the mechanism of this transcriptional repression, we aimed to establish the structural basis of the key intermolecular interaction. In particular, we focused on the analysis of the three-dimensional structure of the complex of Xist and SHARP, and performed analyses using a multidisciplinary approach such as multidimensional NMR, X-ray crystal structure determination, biochemical analysis, and ribo-proteomics. Stable isotope-labeled proteins were prepared and good NMR spectra of the RRM23 moiety were obtained, and the NMR resonance assignments were established. In the NMR measurements of Xist, there were difficulties such as signal loss due to the dynamics between the two repeats. We overcame the problem by partial isotope labeling
Free Research Field	構造生物化学
Academic Significance and Societal Importance of the Research Achievements	遺伝子量補償は染色体全体まるごとの不活化というスケールの大きな発現抑制であり、それを担う複合体は、多くの因子を含み、転写抑制マシナリー全体の分子レベル、構造レベルでの理解が必要である。またxist遺伝子の他の染色体への移植によってトリソミー等の染色体異常による過剰な発現を抑制できることが、細胞レベルの研究では明らかになっており、Xistのもつ染色体全体を不活化する機能を、タンパク質との複合体形成に基づいた分子（構造）レベルまで落とし込んで理解することは、将来の医学的な展開を視野にいれた基礎研究としても、意義深い。本研究はその第１歩となる研究である。