2011-05-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/643310摘要：跟真核生物相同，酵母菌的端粒是由重複並帶有簡單序列的DNA所構成，主要藉由端粒酵素延長。端粒酵素路徑裡許多必需的基因若是有一個被剃除，酵母菌的端粒就會緩慢的縮短，染色體就會持續損失，最終大部分細胞會死亡。然而，仍有一小部份細胞能夠繼續分裂。這些存活株只會由具有重組能力的酵母菌產生。大部分缺少端粒酵素的存活株具有重複subtelomeric片段以及很短的端粒DNA(於Teng et al, 2000, Molecular Cell中稱為第一型存活株)。相對的，第二型存活株僅占總數十分之一，具有很長以及異質化的端粒。人類大部分體細胞並不具有端粒酵素活性，端粒也因此會持續縮短，此現象相似於缺少端粒酵素的酵母菌。相對的，大部分的人類腫瘤以及建構的細胞株都繪表現端粒酵素並且能夠穩定的維持端粒的長度。有一小部份的人類腫瘤以及建構細胞株並不會表現端粒酵素，並具有很長以及異質化的端粒。酵母菌第二型存活株端粒結構就類似這些利用alternative lengthening of telomere (ALT)路徑來延長端粒的細胞。因此，去了解酵母菌第二型存活株重組機制能夠了解高等生物裡不需要端粒酵素而能夠維持端粒的機制。這個研究整體的目標是為了闡明癌細胞端粒重組分子機制並且應用這些發現在癌症治療上。將近20種分子已經被發現會影響端粒重組。在這個部分裡，我們將會專注在Top3以及Sgs1的角色。第一，我們發表資料顯示Sgs1 sumoylation能夠改變缺少端粒酵素活性酵母菌端粒複製。第二，我們最近發現在具有較短端粒的老化細胞裡修復雙股缺口的能力會下降。第三，我們發現TOP2以及TOP3對於端粒重組是必須的。我們會分析在端粒脢缺失的細胞中Sgs1 sumoylation如何改變Top3在端粒重組裡扮演的角色。我們也將會持續尋找TOP3的抑制物。完成這個計畫將會讓治療癌症的方法獲得進展。了解重組機制能夠給予我們具潛力癌症治療方法的暗示。除此之外，這個計劃的結果將會告訴我們有關於在DNA受到損傷要修復時，細胞老化，幹細胞的維持，癌症的形成時整個基因組重新排列所帶有的重大意義。完成所有目標之後這些結果將會對癌症發病以及治療有顯著的影響，並且帶動台灣醫學基因體學的進展。<br> Abstract: In the yeast Saccharomyces, as in most eukaryotes, telomeres consist of a variable amount of simple sequence DNA that is normally synthesized by telomerase. When one of several genes essential for the telomerase pathway is deleted, yeast telomeres slowly shorten, chromosome loss increases, and most cells eventually die. However, a subset of the cells in cultures lacking telomerase continues to divide. These survivors arise only in strains that are recombination proficient. The majority of the cells that survive in the absence of telomerase are multiple tandem copies of the sub-telomeric element and very short tracts of telomeric NA (named type I survivors in Teng et al., 2000, Molecular Cell). In contrast, type II survivors, that comprise about ten percent of the total, have very long and heterogeneous length telomeres. In humans, most somatic cells do not express telomerase, and their telomeres continuously shorten, yielding a phenotype reminiscent of telomerase minus yeast. Conversely, most human tumors and established human cell lines express telomerase and their telomeres stably maintained. A subset of human tumors and established human cell lines do not express telomerase, yet they have exceptionally long and heterogeneous length telomeres. The structure of telomeres in Saccharomyces type II survivors is similar to that of human cell lines that maintain telomeric DNA by this telomerase independent alternative lengthening of telomeres (ALT) pathway. Thus, a mechanistic understanding of the recombination mechanism that yields type II survivors in Saccharomyces may be relevant to telomerase-independent maintenance of telomeres in higher eukaryotes.The overall objective of this study is to elucidate the molecular mechanisms of telomere-telomere recombination in cancer cells and apply further findings to cancer treatments. So far ~20 factors have been identified to modulate telomere recombination. In this grant, we focus on the roles of Top3 and Sgs1. Our published results showed that Sgs1 sumoylation is able to modulate telomere replication in telomerase negative yeast cells. Second, we found that aged cells with shortened telomeres have deficient ability in DNA double-strand break repair. Furthermore, we found that TOP2 and TOP3 are required for telomere recombination. We will study how Sgs1 sumoylation modulates telomere recombination and we will also hunt for small compound inhibitors for TOP3.Accomplishment of the project will advance our approaches to cure cancer. Characterization of telomere recombination has potential therapeutic implications for cancers. Additionally, results from this project will be relevant to questions concerning global genome rearrangement during DNA damage and repair, cellular senescence, stem cell maintenance and cancer formation. The results generated after accomplishing all these goals will have significant impact on the pathogenesis and treatment of cancer, contributing significantly to the progress of genomic medicine.癌症端粒端粒酵素拓樸異構酵素Cancertelomeretelomerasetopoisomerase