c-Myb-C/EBPのベータ協力のメカニズムは,プロモーター上の分離されたサイトからプロモーター上の分離されたサイトへ
Tahir H Tahirov1, Ko Sato, Emi Ichikawa-Iwata
1Kanagawa Academy of Science and Technology, 3-9 Fukuura, Kanazawa-ku, Yokohama 236-0004, Japan. tahir@med.yokohama-cu.ac.jp
Cell
|January 17, 2002
まとめ
細胞髄芽細胞症 (c-Myb) 転写因子は,そのウイルス同位体 (v-Myb) と異なり,C/EBPβと協力して遺伝子発現を調節します. 構造分析は,この機能的差異を可能にする重要な相互作用を明らかにします.
科学分野:
- 分子生物学は分子生物学である.
- 構造生物学 構造生物学とは
- 遺伝子規制 遺伝子規制
背景:
- 細胞性ミエロブラストーシス (c-Myb) と鳥類性ミエロブラストーシスウイルス (AMV) v-Mybは,骨髄性遺伝子調節に関与する転写因子である.
- c-Mybは,v-Mybではないが,C/EBPβと協力して,骨髄型特異遺伝子の転写を活性化する.
- この機能的差異の構造的根拠は不明である.
研究 の 目的:
- c-MybとC/EBP beta.の間の差異的な協力の基礎となる構造的メカニズムを解明する.
- c-Myb/C/EBPベータ複合体とv-Myb/C/EBPベータ複合体の相互作用インターフェースを比較する.
主な方法:
- c-Mybまたはv-MybのDNA結合ドメイン (DBDs),C/EBPβDBD,およびプロモーターDNAを含む複合体の結晶構造を決定した.
- タンパク質とタンパク質の相互作用を確認するためにGSTプルダウンアッセイを使用しました.
- 転写活性を評価するためにルシフェラーゼトランス活性化アッセイを行った.
- 原子力顕微鏡を用いて,DNAのループと長距離相互作用を視覚化しました.
主要な成果:
- 結晶構造は,c-Myb複合体において,DNAに結合したC/EBPβが,別のDNA断片に結合したc-MybのR2領域と相互作用することを明らかにした.
- v-Myb R2領域における点変異は,v-Myb複合体におけるこの相互作用を廃止した.
- GSTプルダウンとルシフェラーゼアッセイは,c-Myb/C/EBPベータ相互作用の機能的重要性を確認した.
- 原子力顕微鏡では,観察された結晶の相互作用が,DNAループを伴うプロモーターの長距離相互作用を模倣することを実証しました.
結論:
- c-MybのR2領域は,C/EBP beta.との協力的な相互作用のために重要である.
- この相互作用により,長距離のDNAループが促進され,骨髄性遺伝子のトランスクリプション制御が強化されます.
- R2の変異によりv-MybがC/EBPβと効果的に相互作用できないことが,その協力の欠如を説明している.
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