Physical and functional interaction between the Y-box binding protein YB-1and human polyomavirus JC virus large T antigen

Y-box binding protein YB-1 is a member of a family of DNA and RNA binding p roteins which have been shown to affect gene expression at both the transcr iptional and translational levels. We have previously shown that YB-1 modul ates transcription from the promoters of the ubiquitous human polyomavirus JC virus (JCV). Here we investigate the physical and functional interplay b etween YB-1 and the viral regulatory protein large T antigen (T-antigen), u sing JCV as a model system. Results of mobility band shift assays demonstra ted that the efficiency of binding of YB-1 to a 23-bp single-stranded viral target sequence was significantly increased when T-antigen was included in the binding reaction mixture. Affinity chromatography and coimmunoprecipit ation assays demonstrated that YB-1 and T-antigen physically interact with each other. Additionally, results of transcription studies demonstrated tha t these two proteins interact functionally on the JCV early and late gene p romoters. Whereas ectopic expression of YB-1 and T-antigen results in syner gistic transactivation of the viral late promoter, YB-1 alleviates T-antige n-mediated transcriptional suppression of the viral early promoter activity . Furthermore, we have localized, through the use of a series of deletion m utants, the sequences of these proteins which are important for their inter action. The T-antigen-interacting region of YB-1 is located in the cold sho ck domain of YB-1 and its immediate banking sequences, and the YB-1-interac ting domain of T-antigen maps to the carboxy-terminal half of T-antigen. Re sults of transient transfection assays with various YB-1 mutants and T-anti gen expression constructs confirm the specificity of the functional interac tion between YB-1 and T-antigen. Taken together, these data demonstrate tha t the cellular factor YB-1 and the viral regulatory protein T-antigen inter act both physically and functionally and that this interaction modulates tr anscription from the JCV promoters.