2021-01-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/654194Argonaute 是一個能與小RNA 結合，並對目標基因的表現進行調控的蛋白。Argonaute蛋白存在大部分的多細胞生物中。雖然過去Argonaute -小RNA 對目標基因的控制多是抑制基因的表現，近年來也開始發現Argonaute -小RNA 對目標基因也會有促進表現的能力。然而，為什麼Argonaute -小RNA 對目標基因同時具有抑制和促進的能力，而又如何決定如何為促進，如何為抑制還未知。線蟲中有26個Argonaute蛋白，而已知這些Argonaute蛋白有明確分工。其中一個名為CSR-1 的Argonaute蛋白在線蟲中是唯一已知會活化基因的Argonaute蛋白。這個計畫的目的在結合遺傳學，結構學，去了解抑制基因及活化基因的Argonaute蛋白的根本不同點的地方，以利將來能更有效地做基因調控 "Argonautes (Agos) are highly conserved proteins that associated with their respected small RNAs, ex, siRNA, miRNA, piRNA, to perform gene regulation of their targeted transcripts. Though most of the siRNA-Ago guided gene regulation are involved in gene silencing, one of C. elegans Agos, CSR-1, is known for its essential role in proper chromosome segregation during cell division. Previous study had shown that by targeting to the nascent transcript, CSR-1 recruit RNA polymerase II to a distinct locus to prevent aberrant transcription on chromatin, which influence the localization of chromosome segregation proteins, CENP-A. The essential role for CSR-1 recruit RNA polymerase II to a distinct locus is not only critical for chromosome segregation, spermiogenesis gene activation and self and non-self gene distinction are both found rely on CSR-1 mediated transcription activation.Moreover, Agos recruit RNA polymerase II to enhance transcription is also observed in human cells. However, what the key structural differences between CSR-1 and other conventional silencing Agos, ex. WAGO-1, is and how the detailed mechanism of CSR-1 recruit RNA polymerase II to the target are unknown. this project will involve in three major aims: Aim 1: Identify essential regions for RNA polymerase II binding site in transcriptional activated AGO, CSR-1. Aim2: Identify structural difference of transcription activated Ago, CSR-1. Aim3: Establishment of in vitro system to reconstitute CSR-1 mediated RNA polymerase II recruitment."基因調控Argonaute gene activation