| 摘要: | 題目：在內質網壓力下APE1對NF-κB活性及NF-κB所調控基因表現之影響。內質網是一個主要用來決定蛋白質要被送至細胞內或細胞外的胞器，主要功能為蛋白質合成、修飾及透過分泌路徑來把蛋白質運送到目的地。當大量累積未正常摺疊的蛋白質所造成的內質網壓力會透過活化ATF6、PERK及Ire1訊息傳遞路徑，另外，許多的轉錄因子如NF-κB、 AP1、 CHOP及ATF4等會參與其中來調節內質網壓力。許多的研究顯示在一些生理狀況或疾病已經被證實跟內質網壓力有關如B細胞分化成漿細胞、腫瘤、病毒感染、退化性神經疾病及第二型糖尿病。研究也顯示內質網壓力可以誘導氧化壓力的產生，當細胞處於氧化壓力下會嚴重干擾到不同特性的蛋白質如轉錄因子NF-κB及造成DNA受損。另一方面，APE1是一個多功能蛋白質並參與DNA修補及調控轉錄因子的氧化還原狀態，APE1現今已知的可以透過調控氧化還原狀態來增強轉錄因子對DNA結合的能力如NF-κB、AP1及p53等。初步結果顯示我們利用反轉錄酶-PCR來分析APE1 mRNA，APE1 mRNA的表現量可以被內質網壓力所誘導，更進一步的發現eIF2α抑制劑可以減弱MCF-7細胞中APE1 mRNA的表現量。同時，內質網壓力下可誘導NF-κB磷酸化及NF-κB結合能力的活性。更進一步，內質網壓力可以誘導p53的表現且我們成功的發現NF-κB參與內質網壓力所誘導p53的表現。在本計畫中我們將著重於三個部份: (一) 分析APE1在內質網壓力所扮演的角色。 (二) 確認在內質網壓力下APE1及NF-κB的關係。 (三) 探討APE1對NF-κB誘導p53表現的影響。我們希望利用內質網壓力的環境來確認APE1及NF-κB活化的關係，這樣的結果可以提供一個重要的資訊應用在腫瘤生成及化學治療方面。總結而論，這樣的機制研究可以提供一個分子基礎來研發具有潛力的臨床用藥。
Topic: The effect of Human apurinic endonuclease 1 (APE1) on NF-κB activity and NF-κB-regulated gene expression under endoplasmic reticulum stress. The endoplasmic reticulum (ER) is a major compartment for proteins destined to the endo/exocytotic pathway. It is responsible for the synthesis, modification and delivery of proteins to their proper target sites within the secretory pathway and the extracellular space. Accumulation of unfolded protein will activate ATF6, PERK and Ire1 signaling pathways in response to ER stress. Otherwise, many transcription factors are involved in regulation of ER stress, such as NF-κB, AP1, CHOP and ATF4. Many studies have shown many physiological conditions or disease were identified in which related to ER stress, e.g. differentiation of B-cells into plasma cells, tumor, viral infection, degenerative neuronal disorders and type II diabetes. Recent studies have shown that reactive oxygen species (ROS) production was increased by ER stress. The oxidative stress will greatly influence the various properties of protein, such transcription factor NF-κB-binding activity and cause DNA damage. On the other hand, Human Apurinic / apyrimidinic endonuclease (APE1) is a multifunctional protein involved in both base excision DNA repair and redox regulation of transcription factors. APE1 is now known to enhance DNA-binding activity of several transcription factors, including NF-κB, AP1, and p53, by regulating their redox states. Our preliminary results indicated that, APE1 mRNA expression level was induced by ER stress as determined with RT-PCR. Furthermore, eIF2α inhibitor could attenuate the induction in MCF-7 cells. Meanwhile, NF-κB phosphorylation and NF-κB-binding activity were also enhanced by ER stress. Moreover, p53 expression was enhanced by ER stress and the induction was probably regulated by NF-κB. In this project, we will focus on three topics: (1) To analyze the role of APE1 in response to ER stress. (2) Identification of relationship between APE1 and NF-κB-binding activity during ER stress. (3) To investigate the effect of APE1 on NF-κB-regulated p53 expression. We hope to identify relationship between APE1 and NF-κB activity or other redox-related transcription factors under ER stress. These results could provide an important insight in tumorgenesis and chemotherapy. Consequently, this mechanistic research may provide a molecular basis to develop a potent pharmaceutical agent in clinical use. |
