碩士 / 國立臺灣大學 / 生化科技學系 / 103 / White spot syndrome virus (WSSV) is a deadly pathogen that causes shrimp death with a mortality rate of 90-100%. So far, twenty-one WSSV immediate-early genes (IE genes) had been found. The protein sequence of the wssv108 IE gene displays 25% identify with Bad/Rrf2 family transcription factor. Previous studies showed that WSSV could annex shrimp transcription factors to activate its own immediate-early genes to get into lytic phase. Recently, LvYY1 (Yin-yang 1) was cloned from Litopenaeus vannamei. A ubiquitously distributed transcription factor YY1 was known to regulate viral immediate-early genes. This study assumed that LvYY1 participates in the pathogenesis of WSSV. Firstly, knockdown of LvYY1 expression in WSSV-infected shrimp caused lower copy numbers of WSSV and delayed cumulative mortality. Moreover, the reporter assay revealed that the promoter activity of wssv108 was activated 3.9 fold by LvYY1 in S2 cell. Deletion analysis of the promoter suggested that 30-mer DNA region (-102 nt/-73 nt) is important to the activation. According to the sequence alignment, this 30-mer DNA region in the wssv108 promoter has no consensus sequence of YY1 binding sites. The electrophoretic mobility shift assay using mutant sequences as competitive oligonucleotides confirmed that recombinant LvYY1 from Sf9 insect cell binds on the sequence from -91 nt to -83 nt of the wssv108 promoter. Furthermore, site-directed mutagenesis of the promoter verified the YY1 binding site is crucial for transcriptional regulation of the wssv108 promoter by LvYY1. Finally, silencing of LvYY1 expression in WSSV-infected shrimp reduced the levels of wssv108 mRNA. Therefore, this study demonstrated that LvYY1 enhances the transactivation activity of wssv108 via an atypical YY1-binding site to affect WSSV infection.
| Identifer | oai:union.ndltd.org:TW/103NTU05106003 |
| Date | January 2015 |
| Creators | Pei-Si Cai, 蔡沛賜 |
| Contributors | Li-Kwan Chang, 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 | 67 |
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