長壽突變果蠅株EP1101的特性研究

Characterization of a longevity mutant EP1101 in Drosophila

許慈恩(Ci-En Xu)

指導教授 ： 汪宏達

國立清華大學/生命科學院/生物科技研究所/碩士(2007年)

https://doi.org/10.6843/NTHU.2007.00409

摘要

在壓力的篩選之下，我們發現數個由P-element所插入到genome的長壽果蠅，有抗壓的能力。其中一個有抗性的突變株是EPEP1101。由壽命長短的測試，EPEP1101在25℃的環境之下，比對照組的果蠅w1118的平均壽命增加34%。使用反轉錄聚合脢連鎖反應法(RT-PCR)在EPEP1101搜尋由EP-element的插入位置附近的目標基因，發現一個轉錄物(transcript)在年輕及老年的果蠅相較於對照組的果蠅有較高的表現。此目標基因是CG33174基因，會同時轉錄出Long form與short form的基因產物。過量表現CG33174於不同的特定組織當中，都發現壽命延長且對氧化的壓力(oxidative stress)有抗性，而且不論用genomic DNA 或cDNA都有相似的作用。根據flybase中對CG33174的序列分析，它具有類解脂脢的功能區(Lipase like domain),進一步分析CG33174有何功能(function)，由三酸甘油脂的測試及測量中性脂肪(Natural lipids)在果蠅冷凍切片的檢體，發現突變株EPEP1101體內脂肪的分布比對照組的果蠅相對的減少。此外，在小鼠中同樣也發現了CG33174的相對基因，想瞭解在不同物種的CG33174相對基因對於壽命長短是否扮演相同的功能，我們表現了CG33174在果蠅全身可增加壽命。另外測試果蠅的心臟能力，用電擊處理果蠅，之後觀察果蠅心臟恢復的情形，發現突變株EP1101心臟恢復的能力比對照組好。本研究對於CG33174基因的增加於果蠅的壽命影響做了一個全面性的探討。

關鍵字

果蠅 ； 氧化壓力 ； 長壽 ； 類解脂脢

並列摘要

Several EP insertion lines have stress resistance was isolated in the stress screening. One of the resistant mutant, EP EP1101 was tested for lifespan, show 34% increase in the mean lifespan than control w1118. To search for the target gene nearby the EP-element insertion site in EP1101, one transcript with higher expression was detected in EP1101 flies compared to those from w1118. The target gene is CG33174 gene, which transcribes two form, long form and short form gene product. We use the different GAL4 line driver to over-express the CG33174 gene of genomic DNA or cDNA in the different tissue of fly. We found that fly have lifespan extension and have resistance to the oxidative stress. In the flybase report, the DNA sequence of CG33174 has a lipase-like domain. CG33174 gene is a novel gene, we want to analyze the gene’s function. A triacylglycerol kit was used to measure the triacylglycerol level in the fly body. The Nile red, a fluorescent dye which combines to the neutral lipid, was used in the fly section. We found that mutant fly EP1101 had decreased level in the lipid contents compared with the control w1118. On the other hand, we wondered whether the CG33174 orthologous gene in different species have the same role in the determination of lifespan extension. I made the construction of Drosophila CG33174 orthologous gene in the mouse to generate the transgenic fly. It shows that the mouse orthologous gene also extends lifespan when expressed in Drosophila. Besides, to test the fly heart performance, we found that the mutant EP1101 have the better ability to restore to normal condition than control fly.

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