本實驗室過去發現arecoline可藉由抑制p53的表現以及其轉錄活性而抑制UV誘導的DNA修復能力。而UV誘導的DNA修復主要透過是核苷酸剪切修復，因此arecoline可能會抑制細胞中的核苷酸剪切修復機制。本論文即是要探討arecoline抑制核苷酸剪切修復可能的分子機制。在參與核苷酸剪切修復的分子中，DDB2主要扮演辨識DNA損傷的角色，XPB與XPD則具有解螺旋活性可打開受損的雙股DNA。本篇研究發現：arecoline處理後，DDB2 啟動子活性及其mRNA表現量受到抑制，但DDB2蛋白質的表現量卻增加，這可能和arecoline影響DDB2蛋白質的降解有關。DDB2的降解如被抑制則可能影響後續核苷酸剪切修復的進行。另外，藉由共同免疫沈澱法及免疫螢光染色的研究發現：arecoline也可能影響p53與XPB及XPD蛋白質之間的交互作用，但此結果對核苷酸剪切修復的影響仍有待進一步的研究。綜合以上結果，本篇論文發現arecoline確實會影響一些核苷酸剪切修復分子，這可能和arecoline抑制DNA修復有密切關係。

Previously our studies showed that arecoline could inhibit DNA repair of UV-induced damages through, at least partly, repressing p53’s expression and transactivation activity. Since UV-induced damages are primarily repaired by the mechanism of nucleotide excision repair (NER), we hypothesized that arecoline might inhibit DNA repair via interfering with the machineries of NER. Among the factors involving in NER, DDB2 plays a role in recognition of DNA lesions. XPB and XPD contain helicase activity and can unwind the damaged double-strand DNA. To explore the molecular mechanism underlying arecoline-mediated inhibition of DNA repair, we examined the effect of arecoline on the promoter activity and expression of DDB2. The results showed that arecoline suppressed DDB2 promoter and mRNA expression. In contrast, the constitutive DDB2 protein level was increased by arecoline treatment, implying that arecoline might affect DDB2 protein stability or its degradation. The degradation of DDB2 is necessary for the downstream repair process of NER. Besides, co-immunoprecipitation and immunofluorescent assay showed that arecoline might affect the protein interaction of p53 and XPB or p53 and XPD. However, the consequences of these arecoline’s effects on NER wait for further investigation. Taken together, this study demonstrated that arecoline indeed could interfere with certain NER genes that might be critical for arecoline-mediated repression of NER.

中文摘要 1
英文摘要 2
一、前言 3
二、研究方向與目的 11
三、研究材料與方法 12
四、結果 34
五、討論 43
六、參考文獻 47
七、結果圖表 53
八、附錄 70
九、附件 75

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