Identification of nucleophosmin/B23, an acidic nucleolar protein, as a stimulatory factor for in vitro replication of adenovirus DNA complexed with viral basic core proteins

The processes governing chromatin remodeling and assembly, which occur prio r to and/or after transcription and replication, are not completely underst ood. To understand the mechanisms of transcription and replication from chr omatin templates, we have established in vitro replication and transcriptio n systems using adenovirus (Ad) DNA complexed with viral basic core protein s, called Ad core, as a template. Using this system, we have previously ide ntified, from HeLa cells, template activating factor-I as a stimulatory fac tor for the Ad core DNA replication. Here, using this system as a tool, we identified and purified a novel template activating factor activity that co nsists of two acidic polypeptides whose apparent molecular masses are 38 kD a and 37 kDa. These two polypeptides correspond to two splicing variants of nucleolar phosphoprotein, nucleophosmin/B23. Recombinant B23 proteins stim ulate the Ad core DNA replication, and the acidic regions of B23 proteins a re important for its activity. In addition, B23 proteins directly bind to c ore histones and transfer them to naked DNA. Furthermore, chromatin compone nts such as histories and topoisomerase II are co-immunoprecipitated with B 23 from cell extracts. These observations lead to a hypothesis that nucleop hosmin/B23 is involved in structural changes of chromatin, thereby regulati ng transcription and replication within the ribosomal DNA region or maintai ning the nucleolar structure. (C) 2001 Academic Press.