磷酸肌醇循環(Phosphoinositide turnover)是一個重要的脂質代謝循環,包含著一些先驅物與訊息傳遞物質的生成以及利用，包括磷酸肌醇(phosphatidylinositol)、甘油二酯(diacylglycerol)、磷脂酰肌醇-4,5-二磷酸(PIP2)。在實驗室先前的研究中，我們發現rdgA以及rdgB這兩個基因的突變都可使果蠅的壽命有顯著的延長，其中rdgA會轉譯出二酰甘油激酶(diacylglycerol kinase)，rdgB 則會轉譯出磷酸肌醇運輸蛋白(phosphatidylinositol transfer protein)，這兩個經由rdgA以及rdgB所轉譯出的蛋白在磷酸肌醇循環中均扮演著重要的角色。因而我們想知道磷酸肌醇循環中其他基因的改變是否也同樣會影響壽命。在我們的研究中，我們已線蟲做為模式生物，並以RNA干擾(RNAi)的方式去抑制磷酸肌醇循環上特定基因的表現量,接著觀測壽命是否有改變。我們發現在抑制pitp-1、egl-8、pll-1、dgk-5以及pld-1的表現量後，線蟲的壽命皆有顯著地延長，而抑制ppk-1與cdgs-1則會明顯地縮短線蟲的壽命，此外我們也發現抑制plpp-1.2以及它的旁系同源基因plpp-1.1與 plpp-1.3的表現量後，均沒有明顯的壽命改變現象。總結來說，在我們的研究中，我們發現磷酸肌醇循環上大部分的基因在表現量被抑制之後,都造成線蟲壽命有顯著的改變,這些證據指出磷酸肌醇循環在生物體壽命的調控上可能具有一定程度的關聯性。

Phosphoinositide turnover is an important lipid metabolism for recycling several precursors and second messengers like phosphatidylinositol (PI), DAG, and PIP2. In our previous study, we find that mutations of rdgA and rdgB which encode diacylglycerol kinase and phosphatidylinositol transfer protein respectively in PI cycle both can extend lifespan in Drosophila. Thus we want to investigate whether other PI cycle genes also play a role in lifespan regulation in C. elegans. In this study we use C. elegans model to explore the effects of PI cycle genes knockdown on C. elegans lifespan through RNA interference. We find that knockdown expression of pitp-1, egl-8, pll-1, dgk-5 and pld-1 can significantly extend lifespan in C. elegans. However, knockdown expression of ppk-1 and cdgs-1 display shortened lifespan. Yet, there is no significant lifespan change in C. elegans upon knockdown of plpp-1.2 and its paralogs plpp-1.1 and plpp-1.3. In summary, our study demonstrates that most PI cycle genes upon knockdown can significantly extend lifespan in C. elegans. Together, our study suggests that PI cycle plays a role in lifespan regulation.

Abstract i
中文摘要 ii
致謝 iii
Table of Contents 1
Introduction 3
Materials and Methods 8
RNA interference 8
C. elegans strains 8
Eggs removing from C. elegans 8
Lifespan measurement 9
RNA extraction 9
Reverse transcription 10
Real-TimeQuantitative RCR 10
Bending test of worms 11
Results 12
Knockdown of dgk-5 extends lifespan in C. elegans, but not by the knockdown of its’ paralogs dgk-2, dgk-3, and dgk-4 12
pitp-1 and Y71G12B.17 knockdown extends lifespan in C. elegans, but not by the knockdown of Y54F10A R.1 12
egl-8 and pll-1 knockdown both extend lifespan in C. elegans, but not by the knockdown of their paralogs 13
pld-1 gene knockdown extends lifespan in C. elegans 13
Knockdown of ppk-1 shortens lifespan in C. elegans 14
Knockdown of cdgs-1 gene shortens lifespan in C. elegans 14
Knockdown of plpp-1.1, plpp-1.2 and plpp-1.3 do not alter lifespan in C. elegans 15
Neuron specific RNAi of pitp-1 can extend lifespan in C. elegans but muscle specific RNAi shows the opposite result 15
DAF-16 activity may not be the primary factor causing the different effect on lifespan between RNAi of egl-8, pll-1 and their paralogs 16
The bag of worms occurs in the RNAi knockdown of plc-1 and plc-3 worms but not in ppk-1 RNAi 16
Discussion 18
Figures 23
Tables 54
Appendix 59
References 64

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