Cloning and Functional Study of Aedes aegypti Domeless (AaDOME) in Mosquito Immunity

• Main Authors: Kai-Ting Yang, 楊凱婷
• Other Authors: Wen-Long Cho
• Format: Others
• Language: zh-TW
• Published: 2008
• Online Access: http://ndltd.ncl.edu.tw/handle/96374687755093027270

碩士 === 國立陽明大學 === 熱帶醫學研究所 === 96 === The JAK/STAT signalling pathway plays important roles in vertebrate development and the regulation of complex cellular processes. Components of the pathway are conserved in Dictyostelium, Caenorhabditis, and Drosophila. The complete sequencing and study of the D. melanogaster genome has been identified as Domeless receptor which involved in JAK/STAT pathway during the development. This project aims to examine the potential role of the Aedes aegypti domeless (AaDOME) in mosquito immunity. The full length AaDOME cDNA has been cloned from A. aegypti. The coding region of AaDOME is 2,743 bp and bear 914 amino acids. Based on sequence analysis, the extra-cellular structure of AaDOME protein has the most similarities to the interleukin 6 (IL-6) receptor family, although its sequence has greatly diverged. Like IL-6 and other interleukin receptors, AaDOME has a cytokine binding homology module (CBM) and three extra-cellular fibronectin-type-III domains (FNIII). Despite having low overall similarity, key amino acids are required for signalling and conserved in the CBM region of AaDOME, which contains a set of four cysteine residues in the N-terminal domain and a WSXWS motif in the C-terminal domain. AaDOME contains these features, but the C-terminal domain of AaDOME has an incomplete WSXWS motif (NTXXXWS). Even though, the sequence similarity of AaDOME compares with the IL-6 receptors is low. The shared characteristics suggest that AaDOME may encode the JAK/STAT receptor in A. aegypti. A preliminary data shows that AaDOME expression is significantly induced by dengue virus infection and in adult female after blood feeding. The coding sequence of the extra-cellular CBM domain was expressed for a recombinant protein which was obtained and purified than produced the polyclonal antibody. The antibody could recognize a protein with 104 kDa in both of A. aegypti adult female and Aag2 cell-line. The study of dsRNA interference (dsRNAi) of AaDOME shows that dsRNA can slightly reduce the expression of downstream AaTEP1 gene which is important in JAK/STAT pathway.