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  • 學位論文

人類DNA拓樸異構酶在基因體損傷及穩定性之功能探討

Study on the Roles of Human Topoisomerases in DNA Damage and Chromosome Stability

指導教授 : 李財坤

摘要


DNA拓樸異構酶在各類生物體都廣泛存在,功能卻又獨特奇妙,隨著演化長河的蛻變,負責起解決伴隨DNA複製、轉錄、重組、修復時應運而生的各式拓樸問題。它們能舒展DNA超螺旋體的緊密結構,也能拆解DNA鎖鏈。作用機制是利用酵素活性中心區域的酪氨酸,與DNA的磷酸根之間形成的磷酸酪氨酸鍵結 (phosphotyrosyl bond),來切割繼而黏合DNA組成份的磷酸二酯鍵 (phosphodiester bond)。人類細胞中,至今發現一共有六種DNA拓樸異構酶,分別為核內的第一型拓樸異構酶、第二型拓樸異構酶α及β同質異構體、第三型拓樸異構酶α及β同質異構體,另外還有粒腺體拓樸異構酶。這些成員生物功能仍未盡知悉,例如最新發現的第三型拓樸異構酶,就還未知功能。另一方面,基於第一型和第二型拓樸異構酶的「DNA剪裁特性」,已經使它們成功被開發為癌症治療藥物的分子標靶。然而關於藥物專一性、療效的探討,與臨床副作用的衍生問題仍待解決。因此在本論文,先於第一章概述拓樸異構酶運行的奇妙宇宙:舉凡DNA超螺旋的起源、調控、維護及平衡,到拓樸異構酶的分類、功能與治療性應用,一一做文獻回顧。接著,我的研究欲進一步對這巧妙的DNA魔術師,掀開它光明與黑暗面的神秘面紗。第二章中,即敘述我們發現了人類第三型拓樸異構酶α同質異構體,具備維護染色質結構暨基因體穩定性的角色。這項發現,開展了探索這位拓樸異構酶家族新成員的生理及抑癌功能之重要面向。我們也在第三章的研究中發現,細胞處理過程會參與活化藉由第二型拓樸異構酶造成的DNA損傷反應。其中DNA複製、轉錄與蛋白酶體,是處理第二型拓樸異構酶造成的DNA損傷必需的三項細胞活動;它們也繼而為此DNA損傷反應開啟的下游反應,像是偵測、修復DNA損傷、或死亡程式所必需。最後,第四章中總結本論文,且提供後續研究之展望。

並列摘要


DNA topoisomerases are ubiquitous, yet magical enzymes that are evolved to solve the topological problems on DNA generated during replication, transcription, recombination and repair. They can relax and/or decatenate DNA by transesterification reaction involving cleavage and religation of DNA phosphodiester bond via active center phosphotyrosyl linkage between DNA and enzyme. In human cells, there are six topoisomerases known to date: nuclear topoisomerase I, topoisomerases IIα and IIβ, topoisomerases IIIα and IIIβ,as well as mitochondria topoisomerase I .The biological roles of many of these members, for example the newly identified topoisomerases IIIα and IIIβ, are still unknown. On the other hand, topoisomerases I and II have been developed as molecular targets of cancer drugs due to their scissoring action on DNA. Nevertheless, many problems remain to be solved regarding the drug selectivity, potency and clinical side effects. In this dissertation, the magic topoisomerase world, including the origin, regulation and maintenance/homeostatic of DNA supercoiling, DNA topoisomerases and their therapeutic implications, is generally reviewed in chapter I. In addition, my studies were intended to expand and offer greater insights into understanding both the bright- and dark-side of this elegant, yet mysterious DNA magician. We uncovered “the roles of human topoisomerase IIIα in chromatin structure and genome stability” in chapter II, which opened up very important aspects about the biological and tumorigenic roles of a young topoisomerase family member. In chapter III, we revealed that “cellular processing pathways contribute to the activation of topoisomerase II-mediated DNA damage responses“. Three cellular activities, replication, transcription and proteasome are required for processing topoisomerase II-linked DNA breakage for initiating downstream DNA damage detection, repair as well as cell death programs. Finally, the conclusion and prospective about this study would be provided in chapter IV.

參考文獻


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[5] J.C. Wang. Cellular roles of DNA topoisomerases: a molecular perspective, Nat Rev Mol Cell Biol 3 (2002) 430-440.

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