The Analysis Of The Wsv249 Promoter And Identification Of Transcription Factor CREB-3

Since it was discovered,due to the high infectiousness and high mortality rate of white spot syndrome virus(WSSV),it has become the main pathogenic factor of the shrimp culture industry,and has seriously affected the development of shrimp industry.The gene expression of WSSV is sequential.Immediate-early genes play a key role in the process of virus infecting host,so this article focuses on analysis of WSSV immediate-early gene wsv249 promoter and the relationships between the transcription factors and promoter.WSSV immediate early(IE)gene wsv249 encodes an E3 ubiquitin ligase that can interact with a shrimp ubiquitin-conjugating enzyme.In this study,to understand the transcriptional regulation of wsv249,a serial of 5'-truncated mutations were made on its promoter and the activities of mutated promoters were analyzed.Four 25 bp regions potentially containing either positive or negative regulatory elements were identified.Notably,the deletion of-275/-250,which abolished a cAMP-response element(CRE),greatly reduced the promoter activity by 84.2%.CRE serves as the binding site for proteins belong to the cAMP responsive element-binding proteins(CREBs)family and the activator protein 1(AP-1)family.Therefore,the regulation of shrimp homolog of c-Jun and CREB on wsv249 promoter was further investigated.We found a CREB homologous protein in shrimp,which is the first discovery of CREB homologous protein by rapid amplification of cDNA ends(RACE).We found that CREB-3 is the most widely distributed in gills,and the lowest in the heart.Through the WSSV virus infection experiment,the transcription of CREB-3 has a rapid increase in gills between 12 and 24 hours after infection.This period corresponds to the expression of immediate-early genes of the WSSV virus.Also,we found that Lvc-Jun greatly upregulated the activity of wsv249 promoter by?12.4 fold,and the CRE at-212/-205 but not the one at-256/-249 was essential for the regulation.In contrast,LvCREB-3 could not activate wsv249 promoter activity.This experiment found a new transcription factors,which contributes to an understanding of relationship between virus and host factors,helping understand viral immediate-early gene transcription regulation,and providing theoretical basis for virus prevention in the future.