Characterization Of The Bovine Foamy Virus 3026 Transcriptional Transactivator Borf1

The genomic nature, replication strategy and the gene expression of foamy viruses(FV) diverge in many aspects from what is commonly accepted as the rules of retrovirus. The transcription of FV genes is dependent on two distinct promoters. A transactivator, termed Tas, is required for transcription from the two viral promoters by interacting with Tas response elements (TREs) in both promoters. Tas also interacts with host cell factors. Nevertheless, little is known about the mechanism of the Tas transactivation.To characterize and define molecular mechanism of Tas transcriptional regulation, the property and functional feature of bovine foamy virus transactivator, known as Borf1, were studied. The unrooted tree of Tas from foamy virus genomes, basic biochemical property and posttranslational modify of Borfl were predicted through bioinformatics analysis. The borfl mRNA pattern at different time point was observed by 5 primer rapid amplification of cDNA ends (5'RACE). After the protein was expressed in BL21 strain of E. coli in a soluble form and purified by 3 kinds of chromatography, its high qualitative antiserum from mouse was prepared and used to analyze its cellular localization. To further explore the mechanism that Borfl regulates gene expression by transcriptional transactivation, its activation domain was mapped by mammalian two-hybrid system and the interaction between Borfl and p300/CBP was detected.It is the first report observed directly that the promoters changed during foamy virus infection. And a cryptic start site of transcription was found, which was located upstream of the internal promoter TATA box and used in the natural infection. Combined with the different splicing patterns of borfl transcript in the different time point, it suggested that there was a posttranscription regulation in the BFV replication. To fulfill its transactivation function, this protein was predominately located in the nuclear and related sequence was between amino acids (aa) 167 to 249. Detected by Gal4 binding domain fusion protein analysis in mammalian cells, the transcriptional activation domain of Borfl was defined to an acidic C-terminal region from amino acids 198 to 249. Importantly, the data from transfection, co-localization and co-immunoprecipitation demonstrated that p300/CBP was involved in the process of Borfl transactivation. Further study is required to complete our understanding on the network mechanism of Borfl gene regulation.