博士 / 國立臺灣大學 / 生化科技學系 / 106 / White spot syndrome virus (WSSV), which causes the shrimp white spot syndrome (WSD), is the most threatening virus to the shrimp aquaculture. WSD leads to the high mortality rate up to 100% in 3 to 10 days after infection. Even now, there is still no effective treatment for WSD. After infection, WSSV undergoes a productive replication state called lytic cycle and expresses the immediate early (IE) genes, early genes, and late genes in order. The IE genes express firstly and regulate downstream viral genes expression. So far, 21 WSSV IE genes were identified, but the exact function of these genes are still unknown. Only the gene function and expression regulation of wssv126, which encodes the possible transcription factor IE1, were most studied. Yin-Yang 1 (YY1) is a multifunctional transcription factor that regulates many key cellular processes, including cell proliferation and differentiation. The YY1 family contains a conserved zincfinger domain, which recognizes a specific DNA sequence. In this study, YY1 from Litopenaeus vannamei (LvYY1) was cloned and analyzed. Furthermore the WSSV IE gene transcription regulation mechanisms, which LvYY1 involves, were studied. According to promoter sequence analysis, this study found two possible conserved YY1 binding sites, which are located in the proximal promoter of ie1. Transient transfection revealed that LvYY1 activates ie1transcription apparently. Moreover, point mutations introduced into one of the conserved YY1-binding sites between positions -119 and -126 in the ie1 promoter represses this capability. Electrophoretic mobility shift assay (EMSA) results further indicated that LvYY1 binds to the conserved YY1-binding site in the region between -119 to -126 in the ie1 promoter. Chromatin immunoprecipitation (ChIP) analysis also confirmed that LvYY1 binds to the ie1 promoter in WSSV-infected shrimp. Additionally, reporter assay results suggested that LvYY1 is involved in basal transcriptional regulation via an interaction with L. vannamei TATA-binding protein, (LvTBP). Knockdown of LvYY1 expression by RNA interference (RNAi) in WSSV-infected shrimp reduced viral replication and viral gene expression significantly. The cumulative mortality of infected shrimp also declined with LvYY1 dsRNA injection. In the present study, the penaeid shrimp YY1 homolog, LvYY1, was reported and analyzed for the first time. The results indicate that WSSV uses host LvYY1 to enhance ie1 expression via an YY1-binding site and the TATA box, thereby facilitating lytic activation and viral replication. These findings are expected to contribute to future studies involving WSSV-host interactions.
| Identifer | oai:union.ndltd.org:TW/106NTU05106005 |
| Date | January 2018 |
| Creators | Ping-Han Huang, 黃柄翰 |
| Contributors | 張麗冠 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 101 |
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