Pcf11蛋白终止转录的N-末端结构域的结构与功能解析

Post transcriptional processing of messenger RNA is essential for the transmission of the genetic information in eukaryotic cells. The Pcf11 protein is an important component of the cleavage/polyadenylation factor IA (CFIA) complex, but it remains structurally not yet fully characterized. Previous studies show that the main function of Pcf11 is to help terminate the RNA polymerase II dependent transcription of mRNA, but the molecular and allosteric mechanism for how Pcf11 dissociates the RNA polymerase II are not clear. In the preliminary work, applicants have newly isolated and identified the solution structure of a conserved region, named Pcf11 2nd NTD (residues 141-233aa); by the experiment in yeast, we have also found that it is essential for yeast cell growth, and required for transcription termination. On the basis of the previous work, we plan firstly to deeply analysis the functional of this domain at the molecular level, then characterize the solution structure details and molecular dynamics for Pcf11(1-233aa) for confirming the complete functional region of transcription termination in Pcf11, we will also identify its allosteric process during transcription termination. Finally, we will investigate the preference RNA sequence for Pcf11(1-233aa) and try to test the possibilities for interventing transcription termination process. All of the above studies can help to interpret the molecular mechanism of how Pcf11 regulates the transcription termination; furthermore, provide the candidate strategy for human intervention transcription termination.

信使RNA的转录后加工对真核细胞遗传信息传递至关重要。Pcf11蛋白是信使RNA前体(pre-mRNA)3’末端加工复合物CFIA的重要组分，其结构仍未完全解析。现有研究提示Pcf11主要功能是协助终止RNA聚合酶II依赖性转录，但Pcf11解离RNA聚合酶II并完成pre-mRNA加工过程的结构变化及机制仍不清楚。申请者在前期工作中鉴定了Pcf11中2nd NTD结构域(141-233aa)的三维结构，并发现该结构域可参与pre-mRNA 3’末端加工。本项目拟深入分析2nd NTD的功能，进一步解析Pcf11的N-末端结构域(1-233aa)的三维结构及其分子动力学，研究其在调节终止转录过程中的构象变化，并在此基础上鉴定其偏好结合的RNA序列，尝试人为干预转录终止过程。本项目的完成，可望明确Pcf11(1-233aa)调节转录终止的结构机理，并为实现人为干预转录终止提供潜在靶点。

信使RNA的转录后加工对真核细胞遗传信息传递至关重要。Pcf11蛋白是信使RNA前体(premRNA)3’末端加工复合物CFIA的重要组分。现有研究提示Pcf11主要功能是协助终止RNA聚合酶II依赖性转录，但Pcf11解离RNA聚合酶II并完成pre-mRNA加工过程的结构变化及机制仍不清楚。申请者在本项目工作中鉴定了Pcf11中2ndNTD结构域(141-233aa)，并解析了其液相核磁共振结构。进一步申请人发现该结构域可参与pre-mRNA 3’末端加工，在此基础上解析了Pcf11的N-末端结构域(1-233aa)的三维结构及其分子动力学，确定了Pcf11_2ndNTD的主要功能是与Pcf11_CID共同调节转录终止。本项目结果补充了Pcf11蛋白所有结构域的三维结构，并在结构生物学角度上补充了Pcf11(1-233aa)调节转录终止的结构机理。