The Analysis Of Human Bocavirus-1(H BoV1) Promoter And The Trans-activati On Of The Non Structural Protein NS1

Human bocavirus1(HBoV1) belongs to the genus Bocavirus within the subfamily parvovirus of the family Parvoviridae, The linear single-stranded genome which has a unique promoter in the5’terminal of genome contains～5.5kb in length with the inverted terminal repeats (ITR) at the both ends. The genome encodes two nonstructural proteins including the NS1and nuclear phosphoprotein NP1as well as two major structural proteins, VP1and VP2. The studies of the NP1have been relatively well documented, such as the presence of an atypical nuclear localization signal of the protein, regulating the expression of inflammatory cytokines, induction of cell apoptosis, and so on. The NS1, as a regulatory protein, was first transcribed and translated post infection, however, little is known about its function. Therefore, it is necessary to investigate the functions and mechanism of this protein during the virus replication.In the present study, we investigated the activity of the HBoV1promoter and the transactivation of the NS1on the promoter. The promoter was truncated from both ends and then inserted into the pGL3-Basic-Vector to find out the core promoter by using dual luciferase report system. We identified some potential transcription factor binding sites from96-145nt important to the activity of HBoV1, the key element was located in the sequence froml46-196nt. To study which nucleotide is essential for promoter activity, some mutations in the key element of the promoter were made between96-145nt. The results showed that the CAAT box, in contrast to the TATA-box/CREB/CdxA, was important for optimum promoter activity. Then we analyzed the nuclear localization signal (NLS) and the nuclear export signal (NES) of the NS1. In the same way, by using dual luciferase report system, we studied the trans-activation of NS1on the promoter and found that the NS1could trans-activate the promoter of AP-1,NF-κB,STAT3, but not GAS. In addition, the amino acids from274-379of the NS1is the key element of trans-activation to HBoV1promoter. Then the full length or truncated NS1was cloned to pEFGP-Nl vector to determine the subcellular localization. As a result, we found that a (or more) nuclear localization signal was located in180-274aa and nuclear export signal in340-562aa.This study demonstrated for the first time that the NS1may play important roles in activation of transcription factors including NF-κB、AP-1and STAT3which are closely related to inflammation cytokines expression, suggesting that the NS1may be involved in HBoV1pathogenesis.