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標題: | 非典型Notch訊息調控埃及斑蚊卵殼黑化之研究 A non-canonical Notch signaling regulates egg melanization in the mosquito Aedes aegypti |
作者: | Chia-Hao Chang 張家豪 |
指導教授: | 蕭信宏(Shin-Hong Shiao) |
關鍵字: | 埃及斑蚊,Notch,JNK,黑化,卵殼過氧化酵素, Aedes aegypti,Notch,JNK,melanization,chorion peroxidase, |
出版年 : | 2014 |
學位: | 碩士 |
摘要: | 蚊子是許多重要傳染疾病的媒介,例如瘧疾、登革熱、黃熱病、西尼羅病毒熱、絲蟲病等。然而其中只有黃熱病已發展出疫苗,而且現有的抗瘧疾藥物也逐漸開始產生抗藥性,因此,藉由控制病媒蚊的數量以減緩疾病傳播的替代性策略是現今努力的目標之一。在蚊子的生活史中,蚊卵產出後需要經過一段時間,卵殼才會黑化及硬化,以保護其內的胚胎。在本實驗室先前的研究中發現,當我們以RNAi的方式抑制埃及斑蚊Notch的表現,會造成部分的卵無法黑化及硬化,此外不論這些卵黑化與否,絕大部分的卵皆無法順利孵化。在哺乳類動物及果蠅的研究中發現Notch是一個高度保留的訊息傳遞路徑,在胚胎發育與決定細胞分化扮演重要的角色。過去的研究證實,典型的Notch訊息傳遞路徑是藉由配體 (ligand)與受體 (receptor) Notch相互結合後,引發Notch被切割,再進一步與下游的CSL共同調控下游基因的表現,而近年的文獻也指出Notch可以在無配體結合的情況下,或是不需要CSL的參與,即可誘發下游基因的表現,但詳細的調控機制至今仍不清楚。因此我們在本研究中想深入探討埃及斑紋Notch調控蚊子生殖能力的分子機制。我們使用Notch訊息路徑的抑制劑來抑制Notch訊息路徑的活化,結果顯示卵殼的黑化與卵的孵化率皆受到明顯抑制,但我們以RNAi的方式分別抑制Notch的配體 (Delta和Serrate) 與下游的轉錄因子CSL,結果卻發現對卵殼黑化無明顯影響。因此我們推測Notch調控卵殼黑化及胚胎發育可能經由非典型的Notch訊息路徑。因此,我們使用JNK的抑制劑來抑制JNK的磷酸化,結果發現卵殼的黑化受到明顯抑制。此外,過去文獻指出,參與卵殼黑化硬化的卵殼過氧化酵素 (chorion peroxidase) 可催化卵殼上酪胺酸 (tyrosine) 的相互鍵結,而我們以JNK抑制劑處理蚊子後,其所產出黑化受到明顯抑制的卵,我們發現其卵殼過氧化酵素的活性有明顯下降的情形。未來我們將進一步釐清Notch調控卵殼黑化及胚胎發育的詳細分子機制,本研究成果將有助於制定蚊媒疾病防治的新策略。 Notch signaling pathway is an evolutionary highly conserved cell-cell signaling pathway. Notch acts as both transmembrane receptor and transcription activator. In canonical pathway, Notch is activated after the binding of ligand to the extracellular domain. This specific binding induces proteolytic cleavage and release of the intracellular domain (Notch intracellular domain, NICD). The binding of NICD to the transcription factor, such as CSL, leads to the activation of target genes. It has been reported that Notch signaling may activate through a ligand or CSL independent (non-canonical) mechanism. However, its detail mechanism remains largely unknown. Our previous results demonstrated that silencing of Notch in the mosquito Aedes aegypti resulted in significant inhibition of egg tanning and hatching rate. In this study, we attempted to further investigate the detail regulation machinery of Notch signaling in the reproduction of Aedes aegypti. To further confirm the role of Notch signaling in egg tanning and embryogenesis, we made use of DAPT, a Notch signaling inhibitor, to examine the effect on mosquito reproduction. To elucidate the role of non-canonical Notch signal in the mosquito reproduction, a non-canonical pathway factor (JNK) was inhibited by SP600125 to examine the effect on egg tanning. Our results showed that inhibition of JNK activity resulted in the reduction of egg tanning and hatching rate. Interestingly, the activity of chorion peroxidase, which catalyzing chorion protein cross-linking during chorion hardening, was also reduced in non-tanning eggs. In the future, we will investigate the non-canonical Notch-mediated mosquito reproduction. Data revealed by this study will be crucial for future study on vector competence and vector control in the field. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/18569 |
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顯示於系所單位: | 微生物學科所 |
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